Student Projects for UMass Chemistry 423 Spring 2011: Difference between revisions

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Instruction and signup page (1-6 below) were posted in Sandbox423 for all students to edit. Student created projects (7-18 below) on separate sandbox pages).
<StructureSection load='1wat' size='400' side='right' scene='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4' caption=''>
 
Instructions and signup page (1-6 below) were posted in Sandbox423 for all students to edit. Students created projects (7-8 below, and additional links in section 3) on separate sandbox pages.


'''This sandbox is in use for UMass Chemistry 423. Others please do not edit this page. Thanks!'''


''' Spring 2011 Chem423 Team Projects: Understanding Drug Mechanisms'''
''' Spring 2011 Chem423 Team Projects: Understanding Drug Mechanisms'''
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{{Clear}}
{{Clear}}
<applet load='1wat' size='[450,338]' frame='true' align='right'
 
caption='Aspartate receptor ligand binding domain (1wat)' scene='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'/>




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John Hickey, Josh Drolet, Josephine Harrington, Andrea Simoni - [[#Influenza M2 Proton Channel|influenza M2 proton channel]]  
John Hickey, Josh Drolet, Josephine Harrington, Andrea Simoni - [[#Influenza M2 Proton Channel|influenza M2 proton channel]]  


[[UMass Chem 423 Student Projects 2011-1]]:


Wednesday 4/27/11
Wednesday 4/27/11


Brittany Forkus, Katie Geldart, Elizabeth Schutsky, Breanna Zerfas - beta adrenergic GPCR [[sandbox226]] [[#Beta Adrenergic GPCR|Beta Adrenergic GPCR]]
Brittany Forkus, Katie Geldart, Elizabeth Schutsky, Breanna Zerfas - Beta Adrenergic GPCR


Lucia Tringali, Shaina Boyle, Jaclyn Somadelis , Dany Mbakop -- HIV Protease [[Sandbox17]]
Lucia Tringali, Shaina Boyle, Jaclyn Somadelis , Dany Mbakop -- HIV Protease  


Andy Kim, Zach Brentzel, Tyler Vlass, Zach Hitzig -- Acetylcholinesterase [[sandbox11]]
Andy Kim, Zach Brentzel, Tyler Vlass, Zach Hitzig -- Acetylcholinesterase




Friday 4/29/11
Friday 4/29/11


Varun Chalupadi, Anthony Laviola, Tiffany Brucker, Alan Stebbins - cyclooxygenase [[sandbox25]]
Varun Chalupadi, Anthony Laviola, Tiffany Brucker, Alan Stebbins - Cyclooxygenase


Inna Brockman, Robert Nathan, Sarena Horava, Nick Cadirov - p38 kinase [[sandbox713]]
[[UMass Chem 423 Student Projects 2011-2]]:


David Peltier, Donald Einck, Ethan Leighton, Chris Coakley - Rituximab Fab [[sandbox99]]
Inna Brockman, Robert Nathan, Sarena Horava, Nick Cadirov - p38 kinase
 
David Peltier, Donald Einck, Ethan Leighton, Chris Coakley - Rituximab Fab




Monday 5/2/11
Monday 5/2/11


Max Moulton, Sally Stras, Jordan Schleeweis, Anh Huynh  -- HIV reverse transcription [[Sandbox77]]
Max Moulton, Sally Stras, Jordan Schleeweis, Anh Huynh  -- HIV reverse transcription  


Chris Brueckner, Daniel Roy, John Clarkson, Justin Srodulski -- Ketamine in binding complex with NMDA receptor [[sandbox42]]
Chris Brueckner, Daniel Roy, John Clarkson, Justin Srodulski -- Ketamine in binding complex with NMDA receptor


Lyes Khendek, Paul Breslin, William Rowley, Joe Perito, Ashley Rivera - G-Quadruplex [[sandbox888]]
Lyes Khendek, Paul Breslin, William Rowley, Joe Perito, Ashley Rivera - G-Quadruplex


==Students looking for group members==
==Students looking for group members==
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=='''Flu Neuraminidase'''==
=='''Flu Neuraminidase'''==


[[Image:neuraminidase-and-tamiflu.jpg|right|300px]]<br />
[[Image:neuraminidase-and-tamiflu.jpg|left|300px]]<br />
{{clear}}
===Introduction===
===Introduction===


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===Overall Structure===
===Overall Structure===
{{STRUCTURE_2hu4 |  PDB=2hu4 |  SCENE=  }}
Influenza (flu) Neuraminidase is a homotetramer, with four identical subunits. Each subunit consists mostly of antiparallel beta sheets and three alpha helices, with a beta-propeller folding pattern (<scene name='Sandbox45/Secondary_structure/2'>Secondary Structure</scene>).
Influenza (flu) Neuraminidase is a homotetramer, with four identical subunits. Each subunit consists mostly of antiparallel beta sheets and three alpha helices, with a beta-propeller folding pattern (<scene name='Sandbox45/Secondary_structure/2'>Secondary Structure</scene>).
As shown, the alpha helices lie toward the center of the protein.  
As shown, the alpha helices lie toward the center of the protein.  
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===Additional Features===
===Additional Features===
{{STRUCTURE_2hu4 |  PDB=2hu4  |  SCENE=  }}
 
Neuraminidase breaks the <scene name='Sandbox45/Sialic_acid_and_neuraminidase/1'>sialic acid</scene> groups from glycoproteins.  <scene name='Sandbox45/Sialic_acid_and_hemagglutinin/1'>Hemagglutinin</scene> from on the virus attaches to the sialic acid groups.  The virus is from the cell and is released when neuraminidase breaks the sialic acid groups.  This will allow flu replication to happen and a neuraminidase inhibitor, such as Tamiflu, is required to stop neuraminidase.  The interaction of Tamiflu with N1 neuraminidase is shown with this <scene name='Avian_Influenza_Neuraminidase,_Tamiflu_and_Relenza/Morph_2hty_to_2hu4/8'>morph</scene> (1).  Tamiflu in this model is bound when it is visible and the loop in moved in towards the drug through induced fit.
Neuraminidase breaks the <scene name='Sandbox45/Sialic_acid_and_neuraminidase/1'>sialic acid</scene> groups from glycoproteins.  <scene name='Sandbox45/Sialic_acid_and_hemagglutinin/1'>Hemagglutinin</scene> from on the virus attaches to the sialic acid groups.  The virus is from the cell and is released when neuraminidase breaks the sialic acid groups.  This will allow flu replication to happen and a neuraminidase inhibitor, such as Tamiflu, is required to stop neuraminidase.  The interaction of Tamiflu with N1 neuraminidase is shown with this <scene name='Avian_Influenza_Neuraminidase,_Tamiflu_and_Relenza/Morph_2hty_to_2hu4/8'>morph</scene> (1).  Tamiflu in this model is bound when it is visible and the loop in moved in towards the drug through induced fit.


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=='''Influenza M2 Proton Channel'''==
=='''Influenza M2 Proton Channel'''==
<Structure load='2KQT' size='500' frame='true' align='right' caption='M2 transmembrane peptide of the influenza A virus in DMPC lipid bilayers bound to deuterated amantadine. PDB: 2KQT' scene='Insert optional scene name here' />


===Introduction===
===Introduction===
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Influenza A, better known as the flu, an infection of the nose, throat, and lungs caused by the influenza virus. Basic symptoms include aches, chills, fever, loss of energy and dizziness, but complications can include pneumonia, encephalitis, bronchitis, and death—about 36,000 people every year die of complications from the flu (CDC).
Influenza A, better known as the flu, an infection of the nose, throat, and lungs caused by the influenza virus. Basic symptoms include aches, chills, fever, loss of energy and dizziness, but complications can include pneumonia, encephalitis, bronchitis, and death—about 36,000 people every year die of complications from the flu (CDC).


The M2 protein is a proton-selective ion channel protein that plays an important role in the life cycle of the influenza A virus. The channel itself is a homotetramer with four identical M2 units, where each M2 protein is a helix stabilized by two disulfide bonds. At a low pH, the channel allows hydrogen ions to enter the viral particle form the endosome, effectively lowering the pH on the interior of the virus. This in turn causes the dissociation of the viral matrix protein M1 from the ribonucleoprotein, a critical step in “uncoating” the virus to introduce its contents to the cytoplasm of the host cell (Stouffer). The M2 protein itself consists of three major protein domains: a stretch of 24 amino acids on the N-terminal end that are exposed to the external environment, 22 (largely hydrophobic) amino acids in the transmembrane region, and 52 amino acids on the C-terminal end which are exposed to the inside of the viral particle (Schnell).   
The <scene name='Student_Projects_for_UMass_Chemistry_423_Spring_2011/Cv/1'>M2 protein is a proton-selective ion channel protein </scene> that plays an important role in the life cycle of the influenza A virus. The channel itself is a homotetramer with four identical M2 units, where each M2 protein is a helix stabilized by two disulfide bonds. At a low pH, the channel allows hydrogen ions to enter the viral particle form the endosome, effectively lowering the pH on the interior of the virus. This in turn causes the dissociation of the viral matrix protein M1 from the ribonucleoprotein, a critical step in “uncoating” the virus to introduce its contents to the cytoplasm of the host cell (Stouffer). The M2 protein itself consists of three major protein domains: a stretch of 24 amino acids on the N-terminal end that are exposed to the external environment, 22 (largely hydrophobic) amino acids in the transmembrane region, and 52 amino acids on the C-terminal end which are exposed to the inside of the viral particle (Schnell).   


The function of the M2 channel can be inhibited by the antiviral drug Amantadine, an inhibition that effectively blocks the virus from taking over the host cell. Amantadine inhibits the replication of influenza A viruses by interfering with the uncoating of the virus within the cell. Amantadine is an M2 inhibitor that blocks the ion channel formed by the M2 protein that spans the viral membrane. By blocking this channel, Amantadine effectively prevents the acidification and subsequent release of viral elements into the host cell. Unfortunately, the M2 gene is susceptible to mutations. When one of five (of the 22) amino acids in the transmembrane region is suitably substituted, Amantadine no longer binds in such a way that would block the motion of the protons into the virus.  As of 2009, the CDC noted that a full 100% of Influenza A viruses of types H3N2 and 2009 pandemic flu samples cultured in the United States showed a resistance to Amantadine (CDC).
The function of the M2 channel can be inhibited by the antiviral drug Amantadine, an inhibition that effectively blocks the virus from taking over the host cell. Amantadine inhibits the replication of influenza A viruses by interfering with the uncoating of the virus within the cell. Amantadine is an M2 inhibitor that blocks the ion channel formed by the M2 protein that spans the viral membrane. By blocking this channel, Amantadine effectively prevents the acidification and subsequent release of viral elements into the host cell. Unfortunately, the M2 gene is susceptible to mutations. When one of five (of the 22) amino acids in the transmembrane region is suitably substituted, Amantadine no longer binds in such a way that would block the motion of the protons into the virus.  As of 2009, the CDC noted that a full 100% of Influenza A viruses of types H3N2 and 2009 pandemic flu samples cultured in the United States showed a resistance to Amantadine (CDC).
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Additional features -- John Hickey
Additional features -- John Hickey
== '''Beta-1 Adrenergic GPCR'''  ==
{{STRUCTURE_2y01 |  PDB=2y01  |  SCENE=  }}
===Introduction===
G-Protein Coupled Receptors (GPCRs) are a type of transmembrane protein that are used to pass extracellular signals into the interior, or cytoplasm, of the cell. The signal is passed to the interior of the cell by a conformational change that occurs in the G-protein on the cytoplasmic side of the cell, due to the binding of a ligand to a receptor on the extracellular side of the cell. The protein is called a  G-protein because its activity depends on the binding of a guanyl nucleotide.
The signaling begins with a ligand  and causes the conformational changes necessary for the G-protein to activate another transmembrane protein called andenylate cyclase. This transmembrane protein is activated by the binding of the guanosine triphosphate (GTP)-bound alpha subunit of the G-protein. The cytoplasmic G-protein is heterotrimeric, meaning that it has <scene name='Sandbox226/G-protein_subunits/1'>three distinct protein domains</scene>: alpha (purple), beta (blue) and gamma (yellow). Upon binding of the extracellular ligand the heterotrimer breaks into the beta-gamma complex and the activated alpha subunit, both of which are membrane anchored. When the GTP binds the alpha subunit the affinity of the beta-gamma complex for the alpha subunit decreases, so these two domains of the G-protein separate. The binding of the activated alpha subunit to the adenylate cyclase protein triggers the formation of cAMP from adenosine triphosphate (ATP), in which cAMP is a mobile signal transducer that can act as an activator of other proteins, often kinases, within the cell upon binding events. The small molecule cAMP is formed for as long as the activated alpha subunit of the G-protein in bound to adenylate. Also, since one ligand binding event can activate multiple G-proteins, and one adenylate cyclase protein can activate the formation of cAMP molecules, the effect of the signal is amplified within the cell.
The alpha subunit is deactivated by a spontaneous hydrolysis reaction that converts GTP into GDP. This allosteric deactivation allows the heterotrimer reform at the base of the GPCR membrane protein. The deactivation process includes the unbinding of the extracellular ligand to return the receptor to its deactivated state, and the dephosphorylation of certain amino acids in the carboxy-terminus tail of the GPCR. The GPCRs can be reactivated when a ligand binds the extracellular receptor again, which is mostly a function of the concentration of ligand in the extracellular space.
There are multiple ligands and small signaling molecules that are utilized in the case of GPCRs. Some of the ligands include epinephrine, norepinephrine, adenosine, and GABA. Some of the small molecules include IP3 and DAG. Each ligand stimulates a different pathway and therefore also calls for the use of second messengers (small molecules) in intracellular signaling.<ref>Berg, Jeremy M.. "Biochemistry." Ed. New York: W.H. Freeman and Company, 2007. Print. pp. 383-388.</ref>
GPCRs are currently the target of many drugs because of their allosteric sites that are separate from direct chemical binding sites of the protein. This allows for conformational changes of the protein without competitive binding at the chemical site. Since binding an allosteric site is reciprocal, the types of cooperativity vary based on the chemistry of the ligands. There are three main advantages to using allosteric binding sites on GPCRs as drug targets: (1) the drug is saturable and therefore can only affect the cells to an upper or lower limit, (2) the response of the cell can change with time and space when using an allosteric modulator, and (3) the potential increased selectivity of the binding site. There are multiple binding sites and ligands that can effect the activity, or non-activity, of the protein. In this case, we are describing the interaction of the drug dobutamine with the <scene name='Sandbox226/Ac1_active_site/1'>beta-1-adrenergic receptor</scene> of a G-protein coupled receptor.
<ref>PMID: 12120504 </ref>
===Overall Structure===
Since this protein is membrane bound, determining the structure was very difficult. Transmembrane proteins are difficult to look at because they are complicated to separate and crystallize. For these reasons, it took about 20 years of studies to determine the structure of β-adrenergic GPCR; the structure was even mutated to increase its thermal stability so that it could be examined<ref>PMID: 21228877</ref>. This protein is made of <scene name='Sandbox226/Two_dimers/1'>two dimers</scene>, each represented in a different color in the scene. Just as is true of most GPCRs, the dimers are each made up of seven <scene name='Sandbox226/Helices_and_ligands/2'> α helices with different ligands</scene>, all of which must span the membrane; the α helices are connected by external and internal loops and are connected in an <scene name='Sandbox226/Antiparallel_representation/1'>anti-parallel</scene> form<ref>PMID: 19458711</ref>. For these α helices to be stable, their middle must be made up of mostly hydrophobic amino acids while their ends are hydrophilic. In this <scene name='Sandbox226/Hydrophobic_and_polar_aas/1'>scene</scene>, hydrophobic amino acids are colored grey while polar amino acids are purple. Though it is shown that some polar amino acids exist on the middle of the helices, they are also mostly on the interior of the helix. This keeps them from being exposed to the lipid membrane and destabilizing the protein. It is also required that all of the hydrogen bonding sites in the α helices are satisfied, so that there are no unfavorable interactions between the lipid inside of the membrane and the protein. As can be seen, the
<scene name='Sandbox226/Hydrogen_bonds/1'>hydrogen bonds</scene> (represented in white) are all between the amino acids in the α helices and not from interactions with the ligand. The ligand to the protein must also have these similar qualities, or else it would not be able exist within the membrane. As seen in this <scene name='Sandbox226/Ligand_ball_and_stick/1'>scene</scene>, the ligand consists of hydrocarbon chains and rings in the center with nitrogen (blue) and oxygen (red) atoms only existing on the ends. This gives the ligands a very similar structure to the membrane, with a hydrophobic center and polar ends. Most of the ligands exist <scene name='Sandbox226/Ligand_with_protein/1'> between the two dimers</scene>, allowing them to participate in binding along with the protein.
<ref name="pdb">[http://www.pdb.org/pdb/explore/explore.do?structureId=2Y01 DOI: 10.2210/pdb2y01/pdb] </ref>
{{STRUCTURE_2y01 |  PDB=2y01  |  SCENE=  }}
===Drug Binding Interactions===
A common technique employed by researchers in the field of modern drug development is to design synthetic ligands that structurally and biochemically mimic natural molecules to induce a desired physiological response. With respect to the beta-adrenergic receptors, several drugs have been created that function to activate or inhibit the receptors for clinical implications such as the treatment of asthma, hypertension, and cardiac dysfunction. In particular, activation of the beta1-adrenergic receptor results in increased cardiovascular output and several drugs including, the total agonists, carmoterol and isoprenaline, and the partial agonists, salbutamol and
<scene name='Sandbox226/Dobutamineee/1'>dobutamine</scene>
, have been designed to take advantage of this feature. The classification as, full agonists, partial agonists, or antagonists is dependent on whether the ligand functions to activate, reduce, or inhibit the natural cellular responses, respectively. However, it is interesting to notice that each of these three classifications has significant effects on the interactions between the ligand and the drug-binding site.
[[Image:Dobutamine_b.png|thumb|Chemical structure of the drug Dobutamine]]
When any of the aforementioned drug agonists bind to the beta-1 receptor, they fit into the natural catecholamine binding-pocket and are capable of inducing three structural changes that result in the protein's overall transition to an active state conformation. The first, and most important, contribution to receptor activation is the
<scene name='Sandbox226/Dobutaminebinding/6'>contraction</scene> of the binding-pocket by 1 angstrom that is stimulated between the alpha carbons of Asn310 and Ser211. Stabilization of the ligand in this pocket is a major factor contributing to ligand efficacy and requires strong hydrogen bonding between the secondary amine and the beta hydroxyl group of the ligand to some of the side chains of protein helices 3 and 7. However, dobutamine lacks the traditional ligand beta-hydroxyl group which, in turn, reduces its potential bonding ability from 4 H-bonds to 2. This weakens the interactions between the helices and is responsible for much of dobutamine’s only partially agonistic behavior.
The second major change that occurs upon agonist binding is the rotamer conformational shift that occurs at
<scene name='Sandbox226/Ser212_conditions/3'>Ser212</scene>. This conformational shift causes Ser212 to form stabilizing hydrogen bonds with Asn310, while simultaneously inducing ligand-mediated bonding interactions between Ser211 and Asn310. These combined actions heighten the extent of interactions between protein helices 5 and 6. The final possible binding change is only observed in total agonists. Full agonists have the ability to additionally induce a conformational change at <scene name='Sandbox226/Ser215-final/2'>Ser215</scene>, which severs the van der waals interactions between Val 172 and Ser215 inducing a subsequent weakening of the interactions between protein helices 4 and 5. The rotameric conformational changes that result from the weakening of interaction between H4 and H5 combined with the strengthening of interactions between H5 and H6, results in a conformational change that is extremely similar to that observed in the familiar and well-studied rhodopsin protein and is believed to significantly contribute to protein activation. It is also important to notice that there are several polar and non-polar interactions that take place right outside of the catecholamine binding-pocket between ligand and protein, which are essential for the specificity of drug binding.
<ref name="pdb" />
===Additional Features===
Adrenergic receptors are the targets of catecholemines, hormones produced in the adrenal glands in response to stress. These ligands result in a wide variety of sympathetic responses depending on the type of ligand and its binding region within the protein<ref name="pdb" />. The alpha and beta domains of the G-protein are broken down into smaller subunits that are more specifically involved in binding.  Different tissues throughout the body often vary in the types of G-protein receptors found in the cells.  Beta 1 adrenergic receptors that bind agonists in the beta 1 site are highly prominent in kidney and heart tissue cells. Beta 2 receptors on the other hand are found in cells in skeletal muscle, lungs, and the GI tract. Beta 3 receptors are found specifically in brown fat cells.  It thus makes sense that introducing a beta 1 binding ligand such as would target heart or kidney cells <ref>Arcangelo V.P.; Peterson A.M. (2006). Pharmacotherapeutics for advanced practice: a practical approach. Lippincott Williams & Wilkins. pp. 205. ISBN 9780781757843. Retrieved 2011-04-21.</ref>.  In the case of dobutamine, the beta 1 site is preferentially bound and results in an increased frequency and intensity of cardiac contractions. Comparatively, beta 2 bound agonists cause smooth muscle and bronchial relaxation and are thus often used in the treatment of asthma. Agonists bound to the alpha receptors can result in vascoconstriction of veins and arteries as well as decreased activity of the smooth muscle in the GI tract <ref>http://www.adrenoceptor.com/betaintro.htm</ref>.
The intensity of a sympathetic response can be controlled with how tightly the agonist is able to bind to its designated receptor. Agonists are broken down into three primary levels of activity; full agonists, partial agonists, and antagonists. Full agonists produce a similar response to that expected from the natural ligand (such as adrenaline or norepinephrine in this case). Dobutamine is considered a partial agonist because it results in a similar but damped response. The primary difference in the interaction of full versus partial agonists is the number of hydrogen bonds formed between the ligand and the protein<ref name="pdb" />. Full agonists form two hydrogen bonds on the 5th alpha helix at Serine 211 and 215 whereas partial agonists only interact with Serine211. <scene name='Sandbox226/Full_agonist_binding/1'>Isoprenaline</scene>, a full agonist, has a similar structure to the native ligand adrenaline and is shown bound to the GPCR. The third category of ligands are called antagonists.  Antagonists inhibit the response of the natural ligand. The term "beta blocker" refers to antagonists that specifically target the beta subunits on adrenergic receptors. Binding of a beta 1 antagonists competitively inhibits the binding of the natural ligand.  This is why beta blockers are used to decrease heart rate to reduce oxygen deprivation of the heart <ref> Khan, M. I. Gabriel (2006). Encyclopedia of Heart Diseases. Elsevier. p. 160. ISBN 0124060616. Retrieved 2010-09-10. </ref>.  <scene name='Sandbox226/Beta_blocker/1'>Cyanopindolol </scene>is shown bound to the beta 1 subunit.  Like the full and partial agonists, this beta blocker binds to Serine 211.  It also hydrogen bonds to Aspartic Acid 121 and Asparagine 329 <ref> http://www.pdb.org/pdb/explore/explore.do?structureId=2VT4 </ref>.
===Credits===
Introduction -- Elizabeth Schutsky
Overall structure -- Breanna Zerfas
Drug binding site -- Brittany Forkus
Additional features -- Katie Geldart
===References===
<references/>
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