https://proteopedia.org/api.php?action=feedcontributions&feedformat=atom&user=Harry+M.+SnyderProteopedia - User contributions [en]2026-09-24T23:41:26ZUser contributionsMediaWiki 1.43.8https://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3658654Sandbox reserved 17522022-11-08T14:05:42Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
<br />
Copy From here down<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<scene name='92/925553/Uvrd/2'>UvrD</scene><br />
<scene name='92/925553/Atp_analog/6'>ATP analog</scene> <br />
Stop here <br />
==Cystic fibrosis transmembrane conductance regulator (CFTR)==<br />
<StructureSection load='5UAK' size='340' side='right' caption='Cystic Fibrosis Transmembrane Conductance regulator' scene=''><br />
The '''CFTR''' is a chloride channel, and is regulated by PKA phosphorylation, cAMP levels, and ATP/ADP ratios. Mutations in the CFTR cause the disease cystic fibrosis.<br />
<br />
CFTR is a mostly <scene name='78/785332/Secondary_structure/1'>alpha helical</scene> protein. The membrane spanning segments can be clearly seen with coloring by <scene name='78/785332/Hydrophobicity/1'>hydrophobicity</scene>, which shows hydrophobic residues in gray and hydrophilic residues in purple.<br />
<br />
The extracellular end of the channel has several <scene name='78/785332/Ec_cl_selection/1'>positively charged</scene> residues that are important for recruiting chloride ions to the channel. A number of <scene name='78/785332/Plus_channel/1'>positively charged</scene> residues line the channel. In the unphosphorylated state (as this structure is), a <scene name='78/785332/Regulatory_domain/2'>regulatory domain</scene> blocks the activity of the channel (the connecting segments are not visible in the structure). It contains several negatively charged residues; when the protein is phosphorylated, this segment is repelled, causing a structural change. <ref>PMID:28340353</ref><br />
<br />
CFTR contains two <scene name='78/785332/Nbd/2'>nucleotide binding domains</scene> (NBD's), which both contain <scene name='78/785332/Walker_motifs/2'>Walker motifs</scene>, flexible loops that bind phosphate groups tightly and are highly conserved among ATP-binding proteins. <br />
<br />
==Mutations in Cystic Fibrosis==<br />
<br />
Cystic fibrosis is characterized by decreased chloride transport, which causes mucus to be thicker and stickier. This leads to a variety of problems, including decreased lung capacity, decreased pancreatic enzyme release into the small intestine, increased rates of lung infections, and infertility.<ref>https://ghr.nlm.nih.gov/condition/cystic-fibrosis</ref> There are a wide assortment of mutations that cause cystic fibrosis, with differing symptom severity. The deletion of <scene name='78/785332/F508/1'>F508</scene> causes the protein to not be properly synthesized, and no expression is seen on the cell surface. Other mutations are found in the NBD's; some of these mutations such as S1255P alter the responsiveness to MgATP, while others such as G551S, G1244E, and G1239D decrease the frequency of channel opening.<br />
<br />
Some of the mutations that lead to cystic fibrosis are due to folding errors. There are <scene name='78/785332/Numbered_bundles/2'>12 transmembrane sequences</scene> in CFTR; they are not sequential in their packing. The presence of <scene name='78/785332/Numbered_bundles_pos_res/1'>hydrophilic, positively charged amino acids</scene> in these transmembrane sequences (shown in red) lead to a folding problem: how do you stabilize them until they can be protected by hydrophobic residues and are no longer exposed to the hydrophobic membrane?<br />
<br />
==3D Printed Physical Model of the CFTR protein==<br />
<br />
Shown below are 3D printed physical models of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein. The backbone model on the left is colored by region, with the transmembrane domain white and the atp-binding and regulatory domains colored red. The backbone model on the right is colored by regional repeat, with the first repeat blue, the second repeat green and the regulatory domain colored red. The models have been designed with embedded magnets to disassemble into the key regions of the structure.<br />
<br />
[[Image:cftr1_centerForBioMolecularModeling.jpg|500px]]<br />
[[Image:cftr2_centerForBioMolecularModeling.jpg|500px]]<br />
<br />
====The MSOE Center for BioMolecular Modeling====<br />
<br />
[[Image:CbmUniversityLogo.jpg | left | 150px]]<br />
<br />
The [http://cbm.msoe.edu MSOE Center for BioMolecular Modeling] uses 3D printing technology to create physical models of protein and molecular structures, making the invisible molecular world more tangible and comprehensible. To view more protein structure models, visit our [http://cbm.msoe.edu/educationalmedia/modelgallery/ Model Gallery].<br />
<br />
<br />
<br />
</StructureSection><br />
<br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=DNA_Repair&diff=3644278DNA Repair2022-10-18T23:40:44Z<p>Harry M. Snyder: </p>
<hr />
<div> The discussion on this page is targeted at genetics students, so familiarity with [[DNA]] structure, [[DNA Replication]] and [[Basics of Protein Structure]] is assumed.<br />
<br />
'''DNA Repair''' is necessary to maintain genome fidelity. Errors in DNA can arise from many different sources. Errors introduced in the replication process are the simplest source. This leads to non-Watson-Crick base pairs and local distortions in the helix. Bases can also be damaged by oxidizing agents, alkylating agents or UV light. This page will discuss different strategies for repairing these types of DNA damage.<br />
<br />
<StructureSection load='B-DNA.pdb' size='350' side='right' scene='' caption=''><br />
<br />
==DNA Mismatch Repair by MutH==<br />
<br />
DNA Mismatch Repair (MMR) occurs when a mismatch of DNA bases occurs during DNA replication that is not corrected by the polymerases. This mismatch can be at a single nucleotide or an insertion or deletion of up to 4 bases. An integral protein in MMR is MutH. MutH is an endonuclease, which means it is an enzyme that can digest DNA in the middle of the sequence. However, it is a weak endonuclease so it will only cause a single-stranded nick upstream or downstream of the damaged daughter strand DNA and not the correct parent strand. This allows it to be re-replicated as the correct sequence by DNA polymerase. Homodimers of MutS and MutL bind the mismatched DNA and create a loop that MutH can bind to. Therefore, MutS and MutL are necessary to recruit MutH to nick the DNA. In order to maintain the correct DNA sequence and repair the damaged portion without mutations, MutH must be able to differentiate the incorrect daughter strand from the correct parent strand. In bacteria, the freshly replicated DNA is hemimethylated, meaning that the parent strand is methylated and the daughter strand has not yet been methylated by methyltransferases. MutH then nicks the phosphodiester bond 5' of a GATC palindrome on the umethylated daughter strand. The GATC palindrome can be upstream or downstream of the damaged DNA site by up to 1000 nucleotides. This allows the damaged strand to be destroyed by exonucleases and re-replicated by DNA polymerase as the correct sequence. <br />
<br />
<br />
=== Structure of MutH ===<br />
MutH has two subdomains, the "N" arm and the "C"arm which is based on the N and C termini of the protein. These arms are arranged in a <scene name='92/925551/Muth_arms/1'>"V" shape</scene>. The N arm contains the catalytic core consisting of the <scene name='92/925551/Dek_motif/2'>DEK motif</scene> and an essential Glu56 residue. The catalytic core is where the endonuclease reaction of hydrolyzing the phosphodiester bond occurs. The DEK motif consists of Asp(D)-X(n)-Glu(E)-X-Lys(K) sequence, which contains the Mg2+ required for nicking the phosphodiester bond. The DEK motif is found in most endonucleases, which highlights its importance in catalyzing the hydrolysis of the phosphodiester bond. <br />
<br />
The <scene name='92/925551/Muth_arms/1'>C-arm</scene> is responsible for base recognition and sequence-specific binding of the DNA. The cleft in the V binds the DNA. The C-term residues help to bind the N-arm and are shown to increase DNA binding in the closed position. This allows it to have the correct shape and chemical interactions to bind the damaged daughter strand DNA substrate and catalyze the hydrolysis reaction in the correct location. <br />
<br />
MutH must be able to correctly recognize the GATC palindrome of the damaged umethylated daughter strand in order to cleave it properly. The secondary structure of Beta sheets 3/9/6 and loop 67 of arm "C" bind the GATC sequence in the major groove of the DNA. The N-arm contacts 6 nucleotides of the cleavage strand in the minor groove of the DNA. Lys45/Asp46 interacts with the phosphate backbone to narrow the minor groove of the DNA. Loop C1 Ser65 H-bonds the nitrogen of Ala67 to stabilize the loop. <scene name='92/925551/Loop_67/5'>Loop 67</scene> (residues 184-190) binds the GATC motif. The G and C are hydrogen bonded by residues Asp184/Glu91 and Lys186/Gly187. Tyr212 bonds N6 the of unmodified adenine and Pro185 interacts with methylated adenine. These specific bonds allow for the recognition of hemimethylated DNA and differentiate the parent strand from the daughter strand. Loop BC Lys48 binds the oxygens of the T’s. The active (catalytic) site on the N arm is Glu56, Asp70, Glu77, and Lys79, this makes up the <scene name='92/925551/Dek_motif/2'>DEK motif</scene>. The carboxylates (Glu/Asp) coordinate two Ca+ ions in the active site. Lys79 links the two arms of MutH and allows for the sequence-specific cutting of DNA. the reaction is catalyzed by Lys79, the 3’ phosphate of DNA that is upstream of the GATC palindrome, and the nearby metal ions to activate water for a <scene name='92/925551/Catalytic_site/3'>nucleophilic attack reaction</scene> to create a single-stranded nick in the daughter strand 5' to the palindrome. Once the nick is created, the damaged daughter strand can be destroyed and re-replicated correctly.<br />
<br />
=== References ===<br />
Ban, C., & Yang, W. (1998). Structural basis for MutH activation in E.coli mismatch repair and relationship of MutH to restriction endonucleases. The EMBO <br />
journal, 17(5), 1526–1534. https://doi.org/10.1093/emboj/17.5.1526<br />
<br />
Lee, J. Y., Chang, J., Joseph, N., Ghirlando, R., Rao, D. N., & Yang, W. (2005). MutH complexed with hemi- and unmethylated DNAs: coupling base recognition <br />
and DNA cleavage. Molecular cell, 20(1), 155–166. https://doi.org/10.1016/j.molcel.2005.08.019<br />
<br />
Voet, D., Voet, J. G., &amp; Pratt, C. W. (2013). Fundamentals of Biochemistry: Life at the molecular level. Wiley.<br />
<br />
<br />
<br />
<br />
==RecA Protein Structure and Function==<br />
<br />
<br />
=== Monomer Structure ===<br />
<br />
<scene name='92/925552/5_monomers/4'>RecA</scene> is one of the many proteins that is involved in recombination cross-over events and during recombination repair in response to single strand DNA breaks. RecA is a rather small monomer protein that can multiplex with itself up to thousands of RecA proteins to associate with ssDNA. The structure of RecA was determined through x-ray crystallography and each monomer contains very distinct structural components. These <scene name='92/925552/Reca_domains/4'>components</scene> are a largely helical 30-residue N-terminal region, a 240-residue α/ß ATPase core, and a 64-residue C-terminal <br />
globular domain.<br />
<br />
<br />
=== Monomer Association ===<br />
<br />
<br />
The process of recruiting new RecA monomers is carried out through an ATP-dependent process. This occurs through the binding of ATP to two adjacent <scene name='92/925552/Atpase_core/5'>α/ß ATPase cores</scene> on subsequent RecA monomers. To properly grow the crystal that was used to determine structure through x-ray crystallography, a non-hydrolyzable analog of ATP is used. This analog has the shorthand formula of ADP-AlF4-Mg. Specifically, the aluminum tetrafluoride is bound to the adenine diphosphate in the ɣ position. Several residues are involved in the hydrolysis of ATP to coordinate strand exchange after binding to ssDNA has occurred. On one of the RecA monomers, two lysine residues, Lys 248 and Lys 250, are responsible for coordinating with the ɣ phosphate stabilizing it. Lys 250 has also been implicated to have an additional function: to coordinate a glutamic acid, Glu 96, on the adjacent RecA monomer. This coordination with Glu 96 is achieved through hydrogen bonding and is believed to be critical for the catalytic mechanism. Specifically, there is a complex network of hydrogen bonding that is occurring between several other residues to rotate Glu 96 to a more favorable conformation enabling Glu 96 to act as a nucleophile.<br />
<br />
<br />
=== ssDNA Binding ===<br />
<br />
<br />
Once several RecA monomers have coordinated with one another, they coordinate with ssDNA to form a repeating structure that contains exactly three nucleotides for every RecA monomer. However, this does not mean that each nucleotide triplet only interacts with a single RecA monomer. In reality, each RecA monomer spans three nucleotides, but the nucleotide triplet interacts with the other two RecA surrounding it in both the 5' and 3' direction. Essentially, each nucleotide triplet is interacting with three different RecA monomers named RecA5', RecA0, and RecA3' based on their relative location to the nucleotide triplet. The first nucleotide of the triplet is bound by both RecA5' and RecA0, the second is bound only by Rec0 and the third is bound by both Rec0 and Rec3'. <scene name='92/925552/Phosphate_interactions/9'>Hydrogen bonding</scene> is responsible for stabilizing ssDNA within this conformation. Specifically, the phosphate backbone of the nucleotide triplet is what interacts with the RecA monomer residues through hydrogen bonding (dashed lines). Interestingly, the hydrogen bonding interactions that are occurring do not always use the side chains, but often will interact with the amide groups on amino acid backbones. For example, the first phosphate group within a nucleotide triplet will interact with the backbone amide of Met 197 from RecA5' and the amide backbone of Asn 123 from RecA0. The second phosphate of the triplet interacts with Gly 211 and Gly 212 on RecA0. The third phosphate of the triplet is unique as it interacts with the side chains of Ser 172 and Arg 176.<br />
<br />
<br />
=== Strand Exchange Mechanism ===<br />
<br />
<br />
Once a RecA filament has properly formed and coordinated with ssDNA, a complementary DNA strand must be located. Once a complementary strand is located, the donor <scene name='92/925552/Strand_exchange_2/4'>dsDNA</scene> is wound into the filament complex where the ssDNA and dsDNA form a temperate three-stranded DNA intermediate. Another protein complex, RecBCD, not modeled here, helps resolve the strand exchange process via the formation of a holiday junction. The process of strand exchange is heavily mediated through traditional Watson-Crick base pairing rules, but also by a few residues located within the RecA filament complex. Specifically, Ser 162 on each RecA monomer contacts the phosphate groups near the nucleotide triplet. Additionally, Met 164 increases the spacing between nucleotide triplets by inserting itself into the gap between them. This insertion allows for more strict base pairing stabilization. The final residue implicated in strand exchange appears to have a proofreading mechanism similar to that of DNA polymerases. This residue is <scene name='92/925552/Arg_169/2'>Arg 169</scene> and has been implicated in having base-pairing proofreading abilities by hydrogen bonding with O2 groups in thymidine bases (DT 7-9). This interaction is able to check for proper Watson-Crick base pairing because the bond lengths associated with proper base pairing will allow for proper interactions between Arg 169 and thymidine. Incorrect base pairing will cause thymidine residues to shift position preventing necessary Arg 169 interactions. This functionality has been shown through the mutation of Arg 169 to Histidine resulting in ultraviolet sensitivity and increased mismatched base pairing. However, this proofreading mechanism is not completely understood and this functionality may be a fragment of Arg 169 being able to interact with subsequent thymidine bases used in the crystalized DNA structure.<br />
<br />
== References ==<br />
<br />
1. Chen, Z., Yang, H., & Pavletich, N. P. (2008). Mechanism of homologous recombination from the RecA-ssDNA/dsDNA structures. Nature, 453(7194), 489–494. https://doi.org/10.1038/nature06971<br />
<br />
2.Voet, D., Voet, J. G., & Pratt, C. W. (2013). Fundamentals of biochemistry : life at the molecular level. Wiley.<br />
<br />
3. Yang, H., Zhou, C., Dhar, A., & Pavletich, N. P. (2020). Mechanism of strand exchange from RecA–DNA synaptic and D-loop structures. Nature, 586(7831), 801–806. https://doi.org/10.1038/s41586-020-2820-9<br />
<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/2'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/6'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
</StructureSection><br />
<br />
<br />
For more structures, please see [[DNA Replication, Repair, and Recombination]]<br />
<br />
==References==<br />
<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644277Sandbox reserved 17522022-10-18T23:40:06Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
<br />
Copy From here down<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<scene name='92/925553/Uvrd/2'>UvrD</scene><br />
<scene name='92/925553/Atp_analog/6'>ATP analog</scene> <br />
Stop here <br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=DNA_Repair&diff=3644276DNA Repair2022-10-18T23:28:59Z<p>Harry M. Snyder: </p>
<hr />
<div> The discussion on this page is targeted at genetics students, so familiarity with [[DNA]] structure, [[DNA Replication]] and [[Basics of Protein Structure]] is assumed.<br />
<br />
'''DNA Repair''' is necessary to maintain genome fidelity. Errors in DNA can arise from many different sources. Errors introduced in the replication process are the simplest source. This leads to non-Watson-Crick base pairs and local distortions in the helix. Bases can also be damaged by oxidizing agents, alkylating agents or UV light. This page will discuss different strategies for repairing these types of DNA damage.<br />
<br />
<StructureSection load='B-DNA.pdb' size='350' side='right' scene='' caption=''><br />
<br />
==DNA Mismatch Repair by MutH==<br />
<br />
DNA Mismatch Repair (MMR) occurs when a mismatch of DNA bases occurs during DNA replication that is not corrected by the polymerases. This mismatch can be at a single nucleotide or an insertion or deletion of up to 4 bases. An integral protein in MMR is MutH. MutH is an endonuclease, which means it is an enzyme that can digest DNA in the middle of the sequence. However, it is a weak endonuclease so it will only cause a single-stranded nick upstream or downstream of the damaged daughter strand DNA and not the correct parent strand. This allows it to be re-replicated as the correct sequence by DNA polymerase. Homodimers of MutS and MutL bind the mismatched DNA and create a loop that MutH can bind to. Therefore, MutS and MutL are necessary to recruit MutH to nick the DNA. In order to maintain the correct DNA sequence and repair the damaged portion without mutations, MutH must be able to differentiate the incorrect daughter strand from the correct parent strand. In bacteria, the freshly replicated DNA is hemimethylated, meaning that the parent strand is methylated and the daughter strand has not yet been methylated by methyltransferases. MutH then nicks the phosphodiester bond 5' of a GATC palindrome on the umethylated daughter strand. The GATC palindrome can be upstream or downstream of the damaged DNA site by up to 1000 nucleotides. This allows the damaged strand to be destroyed by exonucleases and re-replicated by DNA polymerase as the correct sequence. <br />
<br />
<br />
=== Structure of MutH ===<br />
MutH has two subdomains, the "N" arm and the "C"arm which is based on the N and C termini of the protein. These arms are arranged in a <scene name='92/925551/Muth_arms/1'>"V" shape</scene>. The N arm contains the catalytic core consisting of the <scene name='92/925551/Dek_motif/2'>DEK motif</scene> and an essential Glu56 residue. The catalytic core is where the endonuclease reaction of hydrolyzing the phosphodiester bond occurs. The DEK motif consists of Asp(D)-X(n)-Glu(E)-X-Lys(K) sequence, which contains the Mg2+ required for nicking the phosphodiester bond. The DEK motif is found in most endonucleases, which highlights its importance in catalyzing the hydrolysis of the phosphodiester bond. <br />
<br />
The <scene name='92/925551/Muth_arms/1'>C-arm</scene> is responsible for base recognition and sequence-specific binding of the DNA. The cleft in the V binds the DNA. The C-term residues help to bind the N-arm and are shown to increase DNA binding in the closed position. This allows it to have the correct shape and chemical interactions to bind the damaged daughter strand DNA substrate and catalyze the hydrolysis reaction in the correct location. <br />
<br />
MutH must be able to correctly recognize the GATC palindrome of the damaged umethylated daughter strand in order to cleave it properly. The secondary structure of Beta sheets 3/9/6 and loop 67 of arm "C" bind the GATC sequence in the major groove of the DNA. The N-arm contacts 6 nucleotides of the cleavage strand in the minor groove of the DNA. Lys45/Asp46 interacts with the phosphate backbone to narrow the minor groove of the DNA. Loop C1 Ser65 H-bonds the nitrogen of Ala67 to stabilize the loop. <scene name='92/925551/Loop_67/5'>Loop 67</scene> (residues 184-190) binds the GATC motif. The G and C are hydrogen bonded by residues Asp184/Glu91 and Lys186/Gly187. Tyr212 bonds N6 the of unmodified adenine and Pro185 interacts with methylated adenine. These specific bonds allow for the recognition of hemimethylated DNA and differentiate the parent strand from the daughter strand. Loop BC Lys48 binds the oxygens of the T’s. The active (catalytic) site on the N arm is Glu56, Asp70, Glu77, and Lys79, this makes up the <scene name='92/925551/Dek_motif/2'>DEK motif</scene>. The carboxylates (Glu/Asp) coordinate two Ca+ ions in the active site. Lys79 links the two arms of MutH and allows for the sequence-specific cutting of DNA. the reaction is catalyzed by Lys79, the 3’ phosphate of DNA that is upstream of the GATC palindrome, and the nearby metal ions to activate water for a <scene name='92/925551/Catalytic_site/3'>nucleophilic attack reaction</scene> to create a single-stranded nick in the daughter strand 5' to the palindrome. Once the nick is created, the damaged daughter strand can be destroyed and re-replicated correctly.<br />
<br />
=== References ===<br />
Ban, C., & Yang, W. (1998). Structural basis for MutH activation in E.coli mismatch repair and relationship of MutH to restriction endonucleases. The EMBO <br />
journal, 17(5), 1526–1534. https://doi.org/10.1093/emboj/17.5.1526<br />
<br />
Lee, J. Y., Chang, J., Joseph, N., Ghirlando, R., Rao, D. N., & Yang, W. (2005). MutH complexed with hemi- and unmethylated DNAs: coupling base recognition <br />
and DNA cleavage. Molecular cell, 20(1), 155–166. https://doi.org/10.1016/j.molcel.2005.08.019<br />
<br />
Voet, D., Voet, J. G., &amp; Pratt, C. W. (2013). Fundamentals of Biochemistry: Life at the molecular level. Wiley.<br />
<br />
<br />
<br />
<br />
==RecA Protein Structure and Function==<br />
<br />
<br />
=== Monomer Structure ===<br />
<br />
<scene name='92/925552/5_monomers/4'>RecA</scene> is one of the many proteins that is involved in recombination cross-over events and during recombination repair in response to single strand DNA breaks. RecA is a rather small monomer protein that can multiplex with itself up to thousands of RecA proteins to associate with ssDNA. The structure of RecA was determined through x-ray crystallography and each monomer contains very distinct structural components. These <scene name='92/925552/Reca_domains/4'>components</scene> are a largely helical 30-residue N-terminal region, a 240-residue α/ß ATPase core, and a 64-residue C-terminal <br />
globular domain.<br />
<br />
<br />
=== Monomer Association ===<br />
<br />
<br />
The process of recruiting new RecA monomers is carried out through an ATP-dependent process. This occurs through the binding of ATP to two adjacent <scene name='92/925552/Atpase_core/5'>α/ß ATPase cores</scene> on subsequent RecA monomers. To properly grow the crystal that was used to determine structure through x-ray crystallography, a non-hydrolyzable analog of ATP is used. This analog has the shorthand formula of ADP-AlF4-Mg. Specifically, the aluminum tetrafluoride is bound to the adenine diphosphate in the ɣ position. Several residues are involved in the hydrolysis of ATP to coordinate strand exchange after binding to ssDNA has occurred. On one of the RecA monomers, two lysine residues, Lys 248 and Lys 250, are responsible for coordinating with the ɣ phosphate stabilizing it. Lys 250 has also been implicated to have an additional function: to coordinate a glutamic acid, Glu 96, on the adjacent RecA monomer. This coordination with Glu 96 is achieved through hydrogen bonding and is believed to be critical for the catalytic mechanism. Specifically, there is a complex network of hydrogen bonding that is occurring between several other residues to rotate Glu 96 to a more favorable conformation enabling Glu 96 to act as a nucleophile.<br />
<br />
<br />
=== ssDNA Binding ===<br />
<br />
<br />
Once several RecA monomers have coordinated with one another, they coordinate with ssDNA to form a repeating structure that contains exactly three nucleotides for every RecA monomer. However, this does not mean that each nucleotide triplet only interacts with a single RecA monomer. In reality, each RecA monomer spans three nucleotides, but the nucleotide triplet interacts with the other two RecA surrounding it in both the 5' and 3' direction. Essentially, each nucleotide triplet is interacting with three different RecA monomers named RecA5', RecA0, and RecA3' based on their relative location to the nucleotide triplet. The first nucleotide of the triplet is bound by both RecA5' and RecA0, the second is bound only by Rec0 and the third is bound by both Rec0 and Rec3'. <scene name='92/925552/Phosphate_interactions/9'>Hydrogen bonding</scene> is responsible for stabilizing ssDNA within this conformation. Specifically, the phosphate backbone of the nucleotide triplet is what interacts with the RecA monomer residues through hydrogen bonding (dashed lines). Interestingly, the hydrogen bonding interactions that are occurring do not always use the side chains, but often will interact with the amide groups on amino acid backbones. For example, the first phosphate group within a nucleotide triplet will interact with the backbone amide of Met 197 from RecA5' and the amide backbone of Asn 123 from RecA0. The second phosphate of the triplet interacts with Gly 211 and Gly 212 on RecA0. The third phosphate of the triplet is unique as it interacts with the side chains of Ser 172 and Arg 176.<br />
<br />
<br />
=== Strand Exchange Mechanism ===<br />
<br />
<br />
Once a RecA filament has properly formed and coordinated with ssDNA, a complementary DNA strand must be located. Once a complementary strand is located, the donor <scene name='92/925552/Strand_exchange_2/4'>dsDNA</scene> is wound into the filament complex where the ssDNA and dsDNA form a temperate three-stranded DNA intermediate. Another protein complex, RecBCD, not modeled here, helps resolve the strand exchange process via the formation of a holiday junction. The process of strand exchange is heavily mediated through traditional Watson-Crick base pairing rules, but also by a few residues located within the RecA filament complex. Specifically, Ser 162 on each RecA monomer contacts the phosphate groups near the nucleotide triplet. Additionally, Met 164 increases the spacing between nucleotide triplets by inserting itself into the gap between them. This insertion allows for more strict base pairing stabilization. The final residue implicated in strand exchange appears to have a proofreading mechanism similar to that of DNA polymerases. This residue is <scene name='92/925552/Arg_169/2'>Arg 169</scene> and has been implicated in having base-pairing proofreading abilities by hydrogen bonding with O2 groups in thymidine bases (DT 7-9). This interaction is able to check for proper Watson-Crick base pairing because the bond lengths associated with proper base pairing will allow for proper interactions between Arg 169 and thymidine. Incorrect base pairing will cause thymidine residues to shift position preventing necessary Arg 169 interactions. This functionality has been shown through the mutation of Arg 169 to Histidine resulting in ultraviolet sensitivity and increased mismatched base pairing. However, this proofreading mechanism is not completely understood and this functionality may be a fragment of Arg 169 being able to interact with subsequent thymidine bases used in the crystalized DNA structure.<br />
<br />
== References ==<br />
<br />
1. Chen, Z., Yang, H., & Pavletich, N. P. (2008). Mechanism of homologous recombination from the RecA-ssDNA/dsDNA structures. Nature, 453(7194), 489–494. https://doi.org/10.1038/nature06971<br />
<br />
2.Voet, D., Voet, J. G., & Pratt, C. W. (2013). Fundamentals of biochemistry : life at the molecular level. Wiley.<br />
<br />
3. Yang, H., Zhou, C., Dhar, A., & Pavletich, N. P. (2020). Mechanism of strand exchange from RecA–DNA synaptic and D-loop structures. Nature, 586(7831), 801–806. https://doi.org/10.1038/s41586-020-2820-9<br />
<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/6'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
</StructureSection><br />
<br />
<br />
For more structures, please see [[DNA Replication, Repair, and Recombination]]<br />
<br />
==References==<br />
<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644275Sandbox reserved 17522022-10-18T23:27:56Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
<br />
Copy From here down<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
<scene name='92/925553/Atp_analog/6'>ATP analog</scene> <br />
Stop here <br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644274Sandbox reserved 17522022-10-18T23:25:32Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
<br />
Copy From here down<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
<scene name='92/925553/Atp_analog/5'>ATP analog</scene><br />
Stop here <br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=DNA_Repair&diff=3644273DNA Repair2022-10-18T23:18:12Z<p>Harry M. Snyder: </p>
<hr />
<div> The discussion on this page is targeted at genetics students, so familiarity with [[DNA]] structure, [[DNA Replication]] and [[Basics of Protein Structure]] is assumed.<br />
<br />
'''DNA Repair''' is necessary to maintain genome fidelity. Errors in DNA can arise from many different sources. Errors introduced in the replication process are the simplest source. This leads to non-Watson-Crick base pairs and local distortions in the helix. Bases can also be damaged by oxidizing agents, alkylating agents or UV light. This page will discuss different strategies for repairing these types of DNA damage.<br />
<br />
<StructureSection load='B-DNA.pdb' size='350' side='right' scene='' caption=''><br />
<br />
==DNA Mismatch Repair by MutH==<br />
<br />
DNA Mismatch Repair (MMR) occurs when a mismatch of DNA bases occurs during DNA replication that is not corrected by the polymerases. This mismatch can be at a single nucleotide or an insertion or deletion of up to 4 bases. An integral protein in MMR is MutH. MutH is an endonuclease, which means it is an enzyme that can digest DNA in the middle of the sequence. However, it is a weak endonuclease so it will only cause a single-stranded nick upstream or downstream of the damaged daughter strand DNA and not the correct parent strand. This allows it to be re-replicated as the correct sequence by DNA polymerase. Homodimers of MutS and MutL bind the mismatched DNA and create a loop that MutH can bind to. Therefore, MutS and MutL are necessary to recruit MutH to nick the DNA. In order to maintain the correct DNA sequence and repair the damaged portion without mutations, MutH must be able to differentiate the incorrect daughter strand from the correct parent strand. In bacteria, the freshly replicated DNA is hemimethylated, meaning that the parent strand is methylated and the daughter strand has not yet been methylated by methyltransferases. MutH then nicks the phosphodiester bond 5' of a GATC palindrome on the umethylated daughter strand. The GATC palindrome can be upstream or downstream of the damaged DNA site by up to 1000 nucleotides. This allows the damaged strand to be destroyed by exonucleases and re-replicated by DNA polymerase as the correct sequence. <br />
<br />
<br />
=== Structure of MutH ===<br />
MutH has two subdomains, the "N" arm and the "C"arm which is based on the N and C termini of the protein. These arms are arranged in a <scene name='92/925551/Muth_arms/1'>"V" shape</scene>. The N arm contains the catalytic core consisting of the <scene name='92/925551/Dek_motif/2'>DEK motif</scene> and an essential Glu56 residue. The catalytic core is where the endonuclease reaction of hydrolyzing the phosphodiester bond occurs. The DEK motif consists of Asp(D)-X(n)-Glu(E)-X-Lys(K) sequence, which contains the Mg2+ required for nicking the phosphodiester bond. The DEK motif is found in most endonucleases, which highlights its importance in catalyzing the hydrolysis of the phosphodiester bond. <br />
<br />
The <scene name='92/925551/Muth_arms/1'>C-arm</scene> is responsible for base recognition and sequence-specific binding of the DNA. The cleft in the V binds the DNA. The C-term residues help to bind the N-arm and are shown to increase DNA binding in the closed position. This allows it to have the correct shape and chemical interactions to bind the damaged daughter strand DNA substrate and catalyze the hydrolysis reaction in the correct location. <br />
<br />
MutH must be able to correctly recognize the GATC palindrome of the damaged umethylated daughter strand in order to cleave it properly. The secondary structure of Beta sheets 3/9/6 and loop 67 of arm "C" bind the GATC sequence in the major groove of the DNA. The N-arm contacts 6 nucleotides of the cleavage strand in the minor groove of the DNA. Lys45/Asp46 interacts with the phosphate backbone to narrow the minor groove of the DNA. Loop C1 Ser65 H-bonds the nitrogen of Ala67 to stabilize the loop. <scene name='92/925551/Loop_67/5'>Loop 67</scene> (residues 184-190) binds the GATC motif. The G and C are hydrogen bonded by residues Asp184/Glu91 and Lys186/Gly187. Tyr212 bonds N6 the of unmodified adenine and Pro185 interacts with methylated adenine. These specific bonds allow for the recognition of hemimethylated DNA and differentiate the parent strand from the daughter strand. Loop BC Lys48 binds the oxygens of the T’s. The active (catalytic) site on the N arm is Glu56, Asp70, Glu77, and Lys79, this makes up the <scene name='92/925551/Dek_motif/2'>DEK motif</scene>. The carboxylates (Glu/Asp) coordinate two Ca+ ions in the active site. Lys79 links the two arms of MutH and allows for the sequence-specific cutting of DNA. the reaction is catalyzed by Lys79, the 3’ phosphate of DNA that is upstream of the GATC palindrome, and the nearby metal ions to activate water for a <scene name='92/925551/Catalytic_site/3'>nucleophilic attack reaction</scene> to create a single-stranded nick in the daughter strand 5' to the palindrome. Once the nick is created, the damaged daughter strand can be destroyed and re-replicated correctly.<br />
<br />
=== References ===<br />
Ban, C., & Yang, W. (1998). Structural basis for MutH activation in E.coli mismatch repair and relationship of MutH to restriction endonucleases. The EMBO <br />
journal, 17(5), 1526–1534. https://doi.org/10.1093/emboj/17.5.1526<br />
<br />
Lee, J. Y., Chang, J., Joseph, N., Ghirlando, R., Rao, D. N., & Yang, W. (2005). MutH complexed with hemi- and unmethylated DNAs: coupling base recognition <br />
and DNA cleavage. Molecular cell, 20(1), 155–166. https://doi.org/10.1016/j.molcel.2005.08.019<br />
<br />
Voet, D., Voet, J. G., &amp; Pratt, C. W. (2013). Fundamentals of Biochemistry: Life at the molecular level. Wiley.<br />
<br />
<br />
<br />
<br />
==RecA Protein Structure and Function==<br />
<br />
<br />
=== Monomer Structure ===<br />
<br />
<scene name='92/925552/5_monomers/4'>RecA</scene> is one of the many proteins that is involved in recombination cross-over events and during recombination repair in response to single strand DNA breaks. RecA is a rather small monomer protein that can multiplex with itself up to thousands of RecA proteins to associate with ssDNA. The structure of RecA was determined through x-ray crystallography and each monomer contains very distinct structural components. These <scene name='92/925552/Reca_domains/4'>components</scene> are a largely helical 30-residue N-terminal region, a 240-residue α/ß ATPase core, and a 64-residue C-terminal <br />
globular domain.<br />
<br />
<br />
=== Monomer Association ===<br />
<br />
<br />
The process of recruiting new RecA monomers is carried out through an ATP-dependent process. This occurs through the binding of ATP to two adjacent <scene name='92/925552/Atpase_core/5'>α/ß ATPase cores</scene> on subsequent RecA monomers. To properly grow the crystal that was used to determine structure through x-ray crystallography, a non-hydrolyzable analog of ATP is used. This analog has the shorthand formula of ADP-AlF4-Mg. Specifically, the aluminum tetrafluoride is bound to the adenine diphosphate in the ɣ position. Several residues are involved in the hydrolysis of ATP to coordinate strand exchange after binding to ssDNA has occurred. On one of the RecA monomers, two lysine residues, Lys 248 and Lys 250, are responsible for coordinating with the ɣ phosphate stabilizing it. Lys 250 has also been implicated to have an additional function: to coordinate a glutamic acid, Glu 96, on the adjacent RecA monomer. This coordination with Glu 96 is achieved through hydrogen bonding and is believed to be critical for the catalytic mechanism. Specifically, there is a complex network of hydrogen bonding that is occurring between several other residues to rotate Glu 96 to a more favorable conformation enabling Glu 96 to act as a nucleophile.<br />
<br />
<br />
=== ssDNA Binding ===<br />
<br />
<br />
Once several RecA monomers have coordinated with one another, they coordinate with ssDNA to form a repeating structure that contains exactly three nucleotides for every RecA monomer. However, this does not mean that each nucleotide triplet only interacts with a single RecA monomer. In reality, each RecA monomer spans three nucleotides, but the nucleotide triplet interacts with the other two RecA surrounding it in both the 5' and 3' direction. Essentially, each nucleotide triplet is interacting with three different RecA monomers named RecA5', RecA0, and RecA3' based on their relative location to the nucleotide triplet. The first nucleotide of the triplet is bound by both RecA5' and RecA0, the second is bound only by Rec0 and the third is bound by both Rec0 and Rec3'. <scene name='92/925552/Phosphate_interactions/9'>Hydrogen bonding</scene> is responsible for stabilizing ssDNA within this conformation. Specifically, the phosphate backbone of the nucleotide triplet is what interacts with the RecA monomer residues through hydrogen bonding (dashed lines). Interestingly, the hydrogen bonding interactions that are occurring do not always use the side chains, but often will interact with the amide groups on amino acid backbones. For example, the first phosphate group within a nucleotide triplet will interact with the backbone amide of Met 197 from RecA5' and the amide backbone of Asn 123 from RecA0. The second phosphate of the triplet interacts with Gly 211 and Gly 212 on RecA0. The third phosphate of the triplet is unique as it interacts with the side chains of Ser 172 and Arg 176.<br />
<br />
<br />
=== Strand Exchange Mechanism ===<br />
<br />
<br />
Once a RecA filament has properly formed and coordinated with ssDNA, a complementary DNA strand must be located. Once a complementary strand is located, the donor <scene name='92/925552/Strand_exchange_2/4'>dsDNA</scene> is wound into the filament complex where the ssDNA and dsDNA form a temperate three-stranded DNA intermediate. Another protein complex, RecBCD, not modeled here, helps resolve the strand exchange process via the formation of a holiday junction. The process of strand exchange is heavily mediated through traditional Watson-Crick base pairing rules, but also by a few residues located within the RecA filament complex. Specifically, Ser 162 on each RecA monomer contacts the phosphate groups near the nucleotide triplet. Additionally, Met 164 increases the spacing between nucleotide triplets by inserting itself into the gap between them. This insertion allows for more strict base pairing stabilization. The final residue implicated in strand exchange appears to have a proofreading mechanism similar to that of DNA polymerases. This residue is <scene name='92/925552/Arg_169/2'>Arg 169</scene> and has been implicated in having base-pairing proofreading abilities by hydrogen bonding with O2 groups in thymidine bases (DT 7-9). This interaction is able to check for proper Watson-Crick base pairing because the bond lengths associated with proper base pairing will allow for proper interactions between Arg 169 and thymidine. Incorrect base pairing will cause thymidine residues to shift position preventing necessary Arg 169 interactions. This functionality has been shown through the mutation of Arg 169 to Histidine resulting in ultraviolet sensitivity and increased mismatched base pairing. However, this proofreading mechanism is not completely understood and this functionality may be a fragment of Arg 169 being able to interact with subsequent thymidine bases used in the crystalized DNA structure.<br />
<br />
== References ==<br />
<br />
1. Chen, Z., Yang, H., & Pavletich, N. P. (2008). Mechanism of homologous recombination from the RecA-ssDNA/dsDNA structures. Nature, 453(7194), 489–494. https://doi.org/10.1038/nature06971<br />
<br />
2.Voet, D., Voet, J. G., & Pratt, C. W. (2013). Fundamentals of biochemistry : life at the molecular level. Wiley.<br />
<br />
3. Yang, H., Zhou, C., Dhar, A., & Pavletich, N. P. (2020). Mechanism of strand exchange from RecA–DNA synaptic and D-loop structures. Nature, 586(7831), 801–806. https://doi.org/10.1038/s41586-020-2820-9<br />
<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/4'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
</StructureSection><br />
<br />
<br />
For more structures, please see [[DNA Replication, Repair, and Recombination]]<br />
<br />
==References==<br />
<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=DNA_Repair&diff=3644260DNA Repair2022-10-18T19:56:24Z<p>Harry M. Snyder: </p>
<hr />
<div> The discussion on this page is targeted at genetics students, so familiarity with [[DNA]] structure, [[DNA Replication]] and [[Basics of Protein Structure]] is assumed.<br />
<br />
'''DNA Repair''' is necessary to maintain genome fidelity. Errors in DNA can arise from many different sources. Errors introduced in the replication process are the simplest source. This leads to non-Watson-Crick base pairs and local distortions in the helix. Bases can also be damaged by oxidizing agents, alkylating agents or UV light. This page will discuss different strategies for repairing these types of DNA damage.<br />
<br />
<StructureSection load='B-DNA.pdb' size='350' side='right' scene='' caption=''><br />
<br />
==DNA Mismatch Repair by MutH==<br />
<br />
DNA Mismatch Repair (MMR) occurs when a mismatch of DNA bases occurs during DNA replication that is not corrected by the polymerases. This mismatch can be at a single nucleotide or an insertion or deletion of up to 4 bases. An integral protein in MMR is MutH. MutH is an endonuclease, which means it is an enzyme that can digest DNA in the middle of the sequence. However, it is a weak endonuclease so it will only cause a single-stranded nick upstream or downstream of the damaged daughter strand DNA and not the correct parent strand. This allows it to be re-replicated as the correct sequence by DNA polymerase. Homodimers of MutS and MutL bind the mismatched DNA and create a loop that MutH can bind to. Therefore, MutS and MutL are necessary to recruit MutH to nick the DNA. In order to maintain the correct DNA sequence and repair the damaged portion without mutations, MutH must be able to differentiate the incorrect daughter strand from the correct parent strand. In bacteria, the freshly replicated DNA is hemimethylated, meaning that the parent strand is methylated and the daughter strand has not yet been methylated by methyltransferases. MutH then nicks the phosphodiester bond 5' of a GATC palindrome on the umethylated daughter strand. The GATC palindrome can be upstream or downstream of the damaged DNA site by up to 1000 nucleotides. This allows the damaged strand to be destroyed by exonucleases and re-replicated by DNA polymerase as the correct sequence. <br />
<br />
<br />
=== Structure of MutH ===<br />
MutH has two subdomains, the "N" arm and the "C"arm which is based on the N and C termini of the protein. These arms are arranged in a <scene name='92/925551/Muth_arms/1'>"V" shape</scene>. The N arm contains the catalytic core consisting of the <scene name='92/925551/Dek_motif/2'>DEK motif</scene> and an essential Glu56 residue. The catalytic core is where the endonuclease reaction of hydrolyzing the phosphodiester bond occurs. The DEK motif consists of Asp(D)-X(n)-Glu(E)-X-Lys(K) sequence, which contains the Mg2+ required for nicking the phosphodiester bond. The DEK motif is found in most endonucleases, which highlights its importance in catalyzing the hydrolysis of the phosphodiester bond. <br />
<br />
The <scene name='92/925551/Muth_arms/1'>C-arm</scene> is responsible for base recognition and sequence-specific binding of the DNA. The cleft in the V binds the DNA. The C-term residues help to bind the N-arm and are shown to increase DNA binding in the closed position. This allows it to have the correct shape and chemical interactions to bind the damaged daughter strand DNA substrate and catalyze the hydrolysis reaction in the correct location. <br />
<br />
MutH must be able to correctly recognize the GATC palindrome of the damaged umethylated daughter strand in order to cleave it properly. The secondary structure of Beta sheets 3/9/6 and loop 67 of arm "C" bind the GATC sequence in the major groove of the DNA. The N-arm contacts 6 nucleotides of the cleavage strand in the minor groove of the DNA. Lys45/Asp46 interacts with the phosphate backbone to narrow the minor groove of the DNA. Loop C1 Ser65 H-bonds the nitrogen of Ala67 to stabilize the loop. <scene name='92/925551/Loop_67/5'>Loop 67</scene> (residues 184-190) binds the GATC motif. The G and C are hydrogen bonded by residues Asp184/Glu91 and Lys186/Gly187. Tyr212 bonds N6 the of unmodified adenine and Pro185 interacts with methylated adenine. These specific bonds allow for the recognition of hemimethylated DNA and differentiate the parent strand from the daughter strand. Loop BC Lys48 binds the oxygens of the T’s. The active (catalytic) site on the N arm is Glu56, Asp70, Glu77, and Lys79, this makes up the <scene name='92/925551/Dek_motif/2'>DEK motif</scene>. The carboxylates (Glu/Asp) coordinate two Ca+ ions in the active site. Lys79 links the two arms of MutH and allows for the sequence-specific cutting of DNA. the reaction is catalyzed by Lys79, the 3’ phosphate of DNA that is upstream of the GATC palindrome, and the nearby metal ions to activate water for a <scene name='92/925551/Catalytic_site/3'>nucleophilic attack reaction</scene> to create a single-stranded nick in the daughter strand 5' to the palindrome. Once the nick is created, the damaged daughter strand can be destroyed and re-replicated correctly.<br />
<br />
=== References ===<br />
Ban, C., & Yang, W. (1998). Structural basis for MutH activation in E.coli mismatch repair and relationship of MutH to restriction endonucleases. The EMBO <br />
journal, 17(5), 1526–1534. https://doi.org/10.1093/emboj/17.5.1526<br />
<br />
Lee, J. Y., Chang, J., Joseph, N., Ghirlando, R., Rao, D. N., & Yang, W. (2005). MutH complexed with hemi- and unmethylated DNAs: coupling base recognition <br />
and DNA cleavage. Molecular cell, 20(1), 155–166. https://doi.org/10.1016/j.molcel.2005.08.019<br />
<br />
Voet, D., Voet, J. G., &amp; Pratt, C. W. (2013). Fundamentals of Biochemistry: Life at the molecular level. Wiley.<br />
<br />
<br />
<br />
<br />
==RecA Protein Structure and Function==<br />
<br />
<br />
=== Monomer Structure ===<br />
<br />
<scene name='92/925552/5_monomers/4'>RecA</scene> is one of the many proteins that is involved in recombination cross-over events and during recombination repair in response to single strand DNA breaks. RecA is a rather small monomer protein that can multiplex with itself up to thousands of RecA proteins to associate with ssDNA. The structure of RecA was determined through x-ray crystallography and each monomer contains very distinct structural components. These <scene name='92/925552/Reca_domains/4'>components</scene> are a largely helical 30-residue N-terminal region, a 240-residue α/ß ATPase core, and a 64-residue C-terminal <br />
globular domain.<br />
<br />
<br />
=== Monomer Association ===<br />
<br />
<br />
The process of recruiting new RecA monomers is carried out through an ATP-dependent process. This occurs through the binding of ATP to two adjacent <scene name='92/925552/Atpase_core/5'>α/ß ATPase cores</scene> on subsequent RecA monomers. To properly grow the crystal that was used to determine structure through x-ray crystallography, a non-hydrolyzable analog of ATP is used. This analog has the shorthand formula of ADP-AlF4-Mg. Specifically, the aluminum tetrafluoride is bound to the adenine diphosphate in the ɣ position. Several residues are involved in the hydrolysis of ATP to coordinate strand exchange after binding to ssDNA has occurred. On one of the RecA monomers, two lysine residues, Lys 248 and Lys 250, are responsible for coordinating with the ɣ phosphate stabilizing it. Lys 250 has also been implicated to have an additional function: to coordinate a glutamic acid, Glu 96, on the adjacent RecA monomer. This coordination with Glu 96 is achieved through hydrogen bonding and is believed to be critical for the catalytic mechanism. Specifically, there is a complex network of hydrogen bonding that is occurring between several other residues to rotate Glu 96 to a more favorable conformation enabling Glu 96 to act as a nucleophile.<br />
<br />
<br />
=== ssDNA Binding ===<br />
<br />
<br />
Once several RecA monomers have coordinated with one another, they coordinate with ssDNA to form a repeating structure that contains exactly three nucleotides for every RecA monomer. However, this does not mean that each nucleotide triplet only interacts with a single RecA monomer. In reality, each RecA monomer spans three nucleotides, but the nucleotide triplet interacts with the other two RecA surrounding it in both the 5' and 3' direction. Essentially, each nucleotide triplet is interacting with three different RecA monomers named RecA5', RecA0, and RecA3' based on their relative location to the nucleotide triplet. The first nucleotide of the triplet is bound by both RecA5' and RecA0, the second is bound only by Rec0 and the third is bound by both Rec0 and Rec3'. <scene name='92/925552/Phosphate_interactions/9'>Hydrogen bonding</scene> is responsible for stabilizing ssDNA within this conformation. Specifically, the phosphate backbone of the nucleotide triplet is what interacts with the RecA monomer residues through hydrogen bonding (dashed lines). Interestingly, the hydrogen bonding interactions that are occurring do not always use the side chains, but often will interact with the amide groups on amino acid backbones. For example, the first phosphate group within a nucleotide triplet will interact with the backbone amide of Met 197 from RecA5' and the amide backbone of Asn 123 from RecA0. The second phosphate of the triplet interacts with Gly 211 and Gly 212 on RecA0. The third phosphate of the triplet is unique as it interacts with the side chains of Ser 172 and Arg 176.<br />
<br />
<br />
=== Strand Exchange Mechanism ===<br />
<br />
<br />
Once a RecA filament has properly formed and coordinated with ssDNA, a complementary DNA strand must be located. Once a complementary strand is located, the donor <scene name='92/925552/Strand_exchange_2/4'>dsDNA</scene> is wound into the filament complex where the ssDNA and dsDNA form a temperate three-stranded DNA intermediate. Another protein complex, RecBCD, not modeled here, helps resolve the strand exchange process via the formation of a holiday junction. The process of strand exchange is heavily mediated through traditional Watson-Crick base pairing rules, but also by a few residues located within the RecA filament complex. Specifically, Ser 162 on each RecA monomer contacts the phosphate groups near the nucleotide triplet. Additionally, Met 164 increases the spacing between nucleotide triplets by inserting itself into the gap between them. This insertion allows for more strict base pairing stabilization. The final residue implicated in strand exchange appears to have a proofreading mechanism similar to that of DNA polymerases. This residue is <scene name='92/925552/Arg_169/2'>Arg 169</scene> and has been implicated in having base-pairing proofreading abilities by hydrogen bonding with O2 groups in thymidine bases (DT 7-9). This interaction is able to check for proper Watson-Crick base pairing because the bond lengths associated with proper base pairing will allow for proper interactions between Arg 169 and thymidine. Incorrect base pairing will cause thymidine residues to shift position preventing necessary Arg 169 interactions. This functionality has been shown through the mutation of Arg 169 to Histidine resulting in ultraviolet sensitivity and increased mismatched base pairing. However, this proofreading mechanism is not completely understood and this functionality may be a fragment of Arg 169 being able to interact with subsequent thymidine bases used in the crystalized DNA structure.<br />
<br />
== References ==<br />
<br />
1. Chen, Z., Yang, H., & Pavletich, N. P. (2008). Mechanism of homologous recombination from the RecA-ssDNA/dsDNA structures. Nature, 453(7194), 489–494. https://doi.org/10.1038/nature06971<br />
<br />
2.Voet, D., Voet, J. G., & Pratt, C. W. (2013). Fundamentals of biochemistry : life at the molecular level. Wiley.<br />
<br />
3. Yang, H., Zhou, C., Dhar, A., & Pavletich, N. P. (2020). Mechanism of strand exchange from RecA–DNA synaptic and D-loop structures. Nature, 586(7831), 801–806. https://doi.org/10.1038/s41586-020-2820-9<br />
<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
</StructureSection><br />
<br />
For more structures, please see [[DNA Replication, Repair, and Recombination]]<br />
<br />
==References==<br />
<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644258Sandbox reserved 17522022-10-18T19:52:48Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
<br />
Copy From here down<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/2'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
<br />
Stop here <br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644256Sandbox reserved 17522022-10-18T19:46:25Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
<br />
Copy From here down<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
<br />
Stop here <br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644255Sandbox reserved 17522022-10-18T19:42:01Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
<br />
Copy From here down<br />
== UvrD ==<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644254Sandbox reserved 17522022-10-18T19:30:59Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644253Sandbox reserved 17522022-10-18T19:30:14Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase <ref>Voet, D., Voet, J., &amp; Pratt, C. (2015). Fundamentals of Biochemistry: Life at the Molecular Level (4th ed.). Wiley</ref>. <br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644252Sandbox reserved 17522022-10-18T19:22:20Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP Binding <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [2]. <br />
<br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644251Sandbox reserved 17522022-10-18T19:21:59Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [ATP Binding] <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [2]. <br />
<br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644250Sandbox reserved 17522022-10-18T19:20:40Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [2]. <br />
<br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B. Motifs I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644249Sandbox reserved 17522022-10-18T19:18:20Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [2]. <br />
<br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644248Sandbox reserved 17522022-10-18T19:15:37Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [2]. <br />
<br />
<br />
== UvrD Motifs == <br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD. are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
There are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644247Sandbox reserved 17522022-10-18T19:09:21Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Binding">PMID:17190599</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [2]. <br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function ofUvrD <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644246Sandbox reserved 17522022-10-18T19:06:06Z<p>Harry M. Snyder: </p>
<hr />
<div><structuresection load='2IS4' size='340' side='right'caption='[[2IS4]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Binding">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [2]. <br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function ofUvrD <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644128Sandbox reserved 17522022-10-12T01:24:10Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [2]. <br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function ofUvrD <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644127Sandbox reserved 17522022-10-12T01:22:24Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions <ref name="ATP_Binding" />.<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function ofUvrD <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity <ref name="ATP_Binding" />.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP<ref name="ATP_Binding" />. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644126Sandbox reserved 17522022-10-12T01:20:33Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function ofUvrD <ref name="ATP_Binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644125Sandbox reserved 17522022-10-12T01:18:14Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of<ref name="ATP Binding''/>UvrD.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644124Sandbox reserved 17522022-10-12T01:16:44Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved [[motifs]] are important to maintain the function of<ref name="ATP Binding">UvrD.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644123Sandbox reserved 17522022-10-12T01:14:16Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of <ref name="ATP Binding" /><br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644122Sandbox reserved 17522022-10-12T01:11:56Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [[UvrD]] <ref name="ATP binding" />.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644121Sandbox reserved 17522022-10-12T01:11:15Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD <UvrD name="ATP binding" />UvrD.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644120Sandbox reserved 17522022-10-12T01:10:41Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD <ref name="ATP binding" />UvrD.<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644119Sandbox reserved 17522022-10-12T01:09:12Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD <ref name="ATP binding" /><br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644118Sandbox reserved 17522022-10-12T01:08:38Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD <ref name="ATP bindingl" /><br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644117Sandbox reserved 17522022-10-11T14:34:11Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Bindingl">PMID:7482707</ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
<references /></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644116Sandbox reserved 17522022-10-11T14:29:47Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in [[ATP Binding]] <ref name="ATP Binding">PMID:17190599/ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
{{Reflist}}<br />
<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644115Sandbox reserved 17522022-10-11T14:27:52Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [[RasMol]] <ref name="UvrD">PMID:17190599/ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
{{Reflist}}<br />
<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644114Sandbox reserved 17522022-10-11T14:25:37Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [[RasMol]] <ref name="UvrDl">PMID:17190599/ref>. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
{{Reflist}}<br />
<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644113Sandbox reserved 17522022-10-11T14:20:31Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644112Sandbox reserved 17522022-10-11T14:19:35Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycerol molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycerol molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644103Sandbox reserved 17522022-10-11T11:50:17Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644102Sandbox reserved 17522022-10-11T11:47:16Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so UvrD can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is then ready to proceed forward to the next nucleotide [1].<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644101Sandbox reserved 17522022-10-11T11:45:40Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has hydrogen bonding similar to interactions that E566 has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide [1].<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644100Sandbox reserved 17522022-10-11T11:42:07Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog_complete/1'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide [1].<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644099Sandbox reserved 17522022-10-11T11:34:14Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. <scene name='92/925553/Adp_analog/3'>The ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide [1].<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644098Sandbox reserved 17522022-10-11T11:25:17Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/3'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog/2'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide [1].<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644097Sandbox reserved 17522022-10-11T11:17:26Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are<scene name='92/925553/Uvrd_labeled_motifs_complete/3'> 16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/2'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog/2'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide [1].<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644096Sandbox reserved 17522022-10-11T11:13:23Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding [1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [3]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA [2].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions [1].<br />
In total, there are <scene name='92/925553/Uvrd_labeled_motifs_complete/2'>16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD [1].<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/2'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity [1].<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog/2'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP [1]. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide [1].<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644088Sandbox reserved 17522022-10-11T08:58:18Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions. <br />
In total, there are <scene name='92/925553/Uvrd_labeled_motifs_complete/2'>16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/2'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity. This crystal structure induced a 20 degree rotation between domains.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog/2'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide. <br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref><br />
Voet, Voet, &amp; Pratt. (n.d.). Fundamentals biochemistry 4e : Free download, borrow, and streaming. Internet Archive. Retrieved October 11, 2022, from https://archive.org/details/FundamentalsBiochemistry4e</div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644087Sandbox reserved 17522022-10-11T08:56:35Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions. <br />
In total, there are <scene name='92/925553/Uvrd_labeled_motifs_complete/2'>16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/2'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity. This crystal structure induced a 20 degree rotation between domains.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog/2'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP. <br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide. <br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644085Sandbox reserved 17522022-10-11T08:51:26Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with.<ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding.<ref>PMID:17190599</ref> UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage to DNA<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. There are 4 domains that these motifs fit into (not shown). The domains are 1A, 1B, 2A, and 2B.I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. <ref>PMID:17190599</ref> They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions. <ref>PMID:17190599</ref> <br />
In total, there are <scene name='92/925553/Uvrd_labeled_motifs_complete/2'>16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/2'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity. This crystal structure induced a 20 degree rotation between domains.<ref>PMID:17190599</ref><br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog/2'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP. <ref>PMID:17190599</ref><br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide. <ref>PMID:17190599</ref><br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644084Sandbox reserved 17522022-10-11T08:43:46Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from ATP to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with <ref>PMID:17190599</ref> Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding[1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [2]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage in the DNA[3].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions.<br />
In total, there are <scene name='92/925553/Uvrd_labeled_motifs_complete/2'>16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/2'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity. This crystal structure induced a 20 degree rotation between domains.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog/2'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP.<br />
<br />
<br />
<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide.<br />
<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref></div>Harry M. Snyderhttps://proteopedia.org/index.php?title=Sandbox_reserved_1752&diff=3644082Sandbox reserved 17522022-10-11T08:39:37Z<p>Harry M. Snyder: </p>
<hr />
<div><sub><sub></sub></sub>==DNA RECOGNITION BY GAL4: STRUCTURE OF A PROTEIN/DNA COMPLEX==<br />
<StructureSection load='1d66' size='340' side='right'caption='[[1d66]], [[Resolution|resolution]] 2.70&Aring;' scene=''><br />
== Structural highlights ==<br />
<table><tr><td colspan='2'>[[1d66]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Atcc_18824 Atcc 18824]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D66 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D66 FirstGlance]. <br><br />
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CD:CADMIUM+ION'>CD</scene></td></tr><br />
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1d66 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d66 OCA], [https://pdbe.org/1d66 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d66 RCSB], [https://www.ebi.ac.uk/pdbsum/1d66 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d66 ProSAT]</span></td></tr><br />
</table><br />
== Function ==<br />
[[https://www.uniprot.org/uniprot/GAL4_YEAST GAL4_YEAST]] This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which code for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. <br />
== Evolutionary Conservation ==<br />
[[Image:Consurf_key_small.gif|200px|right]]<br />
Check<jmol><br />
<jmolCheckbox><br />
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d66_consurf.spt"</scriptWhenChecked><br />
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked><br />
<text>to colour the structure by Evolutionary Conservation</text><br />
</jmolCheckbox><br />
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d66 ConSurf].<br />
<div style="clear:both"></div><br />
<div style="background-color:#fffaf0;"><br />
== Publication Abstract from PubMed ==<br />
A specific DNA complex of the 65-residue, N-terminal fragment of the yeast transcriptional activator, GAL4, has been analysed at 2.7 A resolution by X-ray crystallography. The protein binds as a dimer to a symmetrical 17-base-pair sequence. A small, Zn(2+)-containing domain recognizes a conserved CCG triplet at each end of the site through direct contacts with the major groove. A short coiled-coil dimerization element imposes 2-fold symmetry. A segment of extended polypeptide chain links the metal-binding module to the dimerization element and specifies the length of the site. The relatively open structure of the complex would allow another protein to bind coordinately with GAL4.<br />
<scene name='92/925553/Dimer_metal_binding_domain/3'>Dimer Metal Binding Domain</scene><br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/2'>Extended Linker</scene><br />
<br />
<scene name='92/925553/Zinc_binding_domain/1'>Zinc Binding Domain <br />
</scene>: Zinc is responsible for maintaining the secondary structure.<br />
<br />
<scene name='92/925553/Alpha-helical_dimerization_ele/4'>Alpha-Helical Dimerization Element</scene><br />
Gal4 has lots of <scene name='92/925553/Basic/1'>basic amino acids</scene> that interact with DNA<br />
<br />
The Upstream activation sequence (<scene name='92/925553/Uas/1'>UAS</scene>) for Gal 4.It only interacts with one strand of DNA sequence. The UAS is a 881 amino acid protein that binds to DNA, has dimerization, and recruits other transcriptional promoters. This needs galactose in order to be open. <br />
<br />
It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. ('''This is what I found from sources)'''<br />
i. I was searching for this, but I kind of struggled to find this when I was looking through the links.<br />
ii. The protein interacts with one strand of double stranded DNA.<br />
<br />
DNA recognition by GAL4: structure of a protein-DNA complex.,Marmorstein R, Carey M, Ptashne M, Harrison SC Nature. 1992 Apr 2;356(6368):408-14. PMID:1557122<ref>PMID:1557122</ref><br />
<br />
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br><br />
</div><br />
<div class="pdbe-citations 1d66" style="background-color:#fffaf0;"></div><br />
<scene name='92/925553/Dimer/1'>Dimer</scene><br />
==See Also==<br />
*[[Gal3-Gal80-Gal4|Gal3-Gal80-Gal4]]<br />
*[[Hydrogen in macromolecular models|Hydrogen in macromolecular models]]<br />
== References ==<br />
<references/><br />
__TOC__<br />
</StructureSection><br />
[[Category: Atcc 18824]]<br />
[[Category: Large Structures]]<br />
[[Category: Carey, M]]<br />
[[Category: Harrison, S C]]<br />
[[Category: Marmorstein, R]]<br />
[[Category: Ptashne, M]]<br />
[[Category: Double helix]]<br />
[[Category: Protein-dna complex]]<br />
[[Category: Transcription-dna complex]]<br />
<br />
== UvrD ==<br />
<br />
<Structure load='2IS4' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' /><br />
<scene name='92/925553/Uvrd/1'>UvrD</scene>, also known as Helicase II, is one of many components responsible in repairing DNA damage. Helicases use energy from nucleoside triphosphate hydrolysis to unwind double helices in metabolic pathways using nucleic acids. ATP molecules are typically used to store energy shared between phosphate groups that gets released when breaking bonds to drive catabolic reactions. <br />
<br />
Helicases were found in the 1970’s to be DNA-dependent ATPases, meaning that they use ATP hydrolysis to complete its interactions with the different types of nucleic acids it comes into contact with [1]. Helicase II, also called UvrD is the founding member of SF1, one group of six superfamiliies used to identify helicases. SF1 and SF2 members share seven conserved sequence motifs that are involved in ATP binding[1]. UvrD is important in replication, recombination, and repair from ultraviolet damage and mismatched base pairs. Nucleotide excision repair in a normal cell is supposed to correct pyrimidine dimers and other DNA lesions when bases are displaced from their normal positions. UvrD pairs up with the UvrABC endonuclease system, which works with UvrD to displace the DNA. This is then repaired by PolI and DNA ligase [2]. A sub pathway of nucleotide excision repair is transcription-coupled repair, which works with an RNA polymerase to make repairs to damage in the DNA[3].<br />
<br />
<br />
== UvrD Motifs == <br />
There are 7 sequence motifs and a Q motif conserved among the SF1 and the SF2. I, Ia, II-VI are involved in ATP binding. Motifs Ia, III, and V are involved in ssDNA binding. Motif IV is reported to be unique in SF1. They found in their paper, seven new sequence motifs conserved among UvrD homologs. They are Ib, Ic, Id, IVb, IVc, Va, and VIa. These conserved residues are involved in DNA binding or domain 1B and 2B interactions.<br />
In total, there are <scene name='92/925553/Uvrd_labeled_motifs_complete/2'>16 binding motifs</scene> for UvrD, which are conserved in other homologous structures, such as PcrA, Rep, and Srs2. The homologous structures mentioned are Helicase 2 homologs, which appear in different species. These conserved motifs are important to maintain the function of UvrD.<br />
== UvrD Binding Site for ATP analog (AMPPNP) ==<br />
When determining the structure of UvrD, an ATP analog was used. They used an <scene name='92/925553/Atp_analog/2'>ATP analog</scene> so that the last phosphate can't be cleaved. Using the unhydrolyzable analog is beneficial in locking in the structure to observe.The green ion shown in the ATP analog scene is a Mg<sup>2+</sup> ion, which is essential for ATP hydrolysis and interacts with the β and γ phosphates. The magnesium ion is surrounded by essential residues that when altered, have been shown to have reduced ATPase activity. This crystal structure induced a 20 degree rotation between domains.<br />
== UvrD Binding Site for ATP analog (ADP•MgF<sub>3</sub>) ==<br />
To capture the UvrD-DNA-ADP complex, a new crystal structure used ADP•MgF<sub>3</sub> after NaF was added to help improve crystal growth. This structure is believed to be a more authentic transition state analog, which differs from the AMPPNP analog slightly. The <scene name='92/925553/Adp_analog/2'>ADP analog</scene> has a <scene name='92/925553/Adp_e566_and_gol/2'>GOL region</scene>, which is a glycogen molecule, which has similar binding that the DNA backbone has to a 3' OH of the ribose. The DNA isn't actually bound in the crystal structure, but can be used as a model to visualize what hydrogen bonding might look like when connected to the backbone in DNA. <scene name='92/925553/Adp_e566_and_gol_hbonding_comp/2'>This glycogen molecule hydrogen bonds with E566</scene>, which typically would bind to the 3' OH of the ribose of DNA. Another residue, R37 (Not Shown), binds to the 2' OH of ribose, which has weaker hydrogen bonding. This is a structural component that allows UvrD to bind both ATP and dATP.<br />
<br />
<br />
<br />
== Separation Pin ==<br />
The "<scene name='92/925553/Pin_complex/1'>separation pin</scene>" is a part of the 2B domain and is responsible for unwinding the DNA. This uses a 2 step power stroke, one stroke when ATP is bound and another stroke when ADP and P<sub>i</sub> are released. The GIG motif and separation pin work together to unwind the DNA and move it out of the way so it can unwind more DNA. The separation pin also prevents ssDNA once unwound from moving backwards and from reannealing. The proposed method is called the wrench-and-inchworm method, which is when the enzyme binds DNA and attaches at different points and then moves 1 nucleotide per ATP molecule.After an ATP molecule is released, UvrD is ready to proceed forward to the next nucleotide.<br />
<br />
== References ==<br />
{{Reflist}}<br />
<ref>PMID:17190599</ref><br />
<ref>PMID:24402227</ref></div>Harry M. Snyder