https://proteopedia.org/api.php?action=feedcontributions&feedformat=atom&user=Nathan+ClarkeProteopedia - User contributions [en]2026-09-15T09:21:31ZUser contributionsMediaWiki 1.43.8https://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230737Ribonuclease A Catalysis2011-04-15T21:04:49Z<p>Nathan Clarke: </p>
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{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. This ability to alter the pKa of certain residues such as histidines, increases the diversity of reactions that an enzyme can perform.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand <ref name="Wlodrawer1988" />. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Threonine 45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands <ref name="Wlodrawer1988" />. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold <ref name="Wlodrawer1988" />. <br />
<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent <ref name="Raines1998" />.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate <ref name="Raines1998" />. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group <ref name="Raines1998" />.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases <ref name="Vicentini1996" />.<br />
[[Image:RI.PNG|300px|Right|thumb|Figure III: Ribonuclease Inhibitor-RNase A Complex. Left, Ribonuclease Inhibitor (RI)is composed of alternating alpha helix (blue) and beta sheets (green). Right, RI-RNase A inhibition forms when RI complex with the active site cleft of RNase (yellow). Figure generated via ''Pymol'']]<br />
The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein <ref name="Turcotte2008" />. <br />
<br />
<br />
=References=<br />
<br />
{{Reflist| refs=<br />
<ref name="Raines1998">Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref><br />
<ref name="Turcotte2008">Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414</ref> <br />
<ref name="Vicentini1996"> Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834</ref><br />
<ref name="Wlodrawer1988">Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref> <br />
}}<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=File:RI.PNG&diff=1230736File:RI.PNG2011-04-15T21:01:48Z<p>Nathan Clarke: </p>
<hr />
<div></div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230735Ribonuclease A Catalysis2011-04-15T21:01:12Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. This ability to alter the pKa of certain residues such as histidines, increases the diversity of reactions that an enzyme can perform.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand <ref name="Wlodrawer1988" />. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Threonine 45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands <ref name="Wlodrawer1988" />. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold <ref name="Wlodrawer1988" />. <br />
<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent <ref name="Raines1998" />.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate <ref name="Raines1998" />. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group <ref name="Raines1998" />.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases <ref name="Vicentini1996" />. The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein <ref name="Turcotte2008" />. <br />
<br />
<br />
<br />
=References=<br />
<br />
{{Reflist| refs=<br />
<ref name="Raines1998">Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref><br />
<ref name="Turcotte2008">Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414</ref> <br />
<ref name="Vicentini1996"> Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834</ref><br />
<ref name="Wlodrawer1988">Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref> <br />
}}<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230570Ribonuclease A Catalysis2011-04-15T02:25:20Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. This ability to alter the pKa of certain residues such as histidines, increases the diversity of reactions that an enzyme can perform.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Threonine 45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases <ref>Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834</ref>. The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein <ref>Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414</ref>. <br />
<br />
=References=<br />
{{Reflist}}<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230569Ribonuclease A Catalysis2011-04-15T02:20:02Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. This ability to alter the pKa of certain residues such as histidines, increases the diversity of reactions that an enzyme can perform.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Threonine 45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases <ref>Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834</ref>. The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein <ref>Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414</ref>. <br />
<br />
=References=<br />
<br />
1. Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414<br />
<br />
3. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834<br />
<br />
4. Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230567Ribonuclease A Catalysis2011-04-15T02:17:52Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. This ability to alter the pKa of certain residues such as histidines, increases the diversity of reactions that an enzyme can perform.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases <ref>Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834</ref>. The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein <ref>Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414</ref>. <br />
<br />
=References=<br />
<br />
1. Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414<br />
<br />
3. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834<br />
<br />
4. Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230564Ribonuclease A Catalysis2011-04-15T02:11:53Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold <ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</ref>. <br />
<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases <ref>Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834</ref>. The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein <ref>Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414</ref>. <br />
<br />
=References=<br />
<br />
1. Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414<br />
<br />
3. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834<br />
<br />
4. Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230562Ribonuclease A Catalysis2011-04-15T02:07:27Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent <ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068</ref>.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases. The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein 3. <br />
<br />
=References=<br />
<br />
1. Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414<br />
<br />
3. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834<br />
<br />
4. Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230559Ribonuclease A Catalysis2011-04-15T02:03:56Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases. The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein 3. <br />
<br />
=References=<br />
<br />
1. </ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. </ref>Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414<br />
<br />
3. </ref>Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834<br />
<br />
4. </ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230556Ribonuclease A Catalysis2011-04-15T02:02:13Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent <ref>Raines, R.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. RI is a large 50 kD protein that is composed of 16 repeating alpha and beta chains, giving it a noticable horseshoe like appearance. It has been suggested that RI has the highest protein-protein interactions with an approximate dissociation constant (Kd) of 5.8 X 10-14 for almost all types of RNases. The ability to be selective for almost all types of RNases, and yet retain such a high Kd is product of its mechanism. The interior residues of the horseshoe shaped RI are able to bind to the charged residues of the active site cleft of RNase A, such as Lys7, Lys9, Lys 41 and Gln11. By studying the amphibian RNase, Onconase, the residues Lys7 and Gln11 of RNase A were shown to be the most important in this interaction. In onconase, these residues are replaced with non-charged amino acids, which help prevent the binding of RI to the protein 3. <br />
<br />
=References=<br />
<br />
1. </ref>Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. </ref>Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414<br />
<br />
3. </ref>Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834<br />
<br />
4. </ref>Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230529Ribonuclease A Catalysis2011-04-15T01:34:14Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor<br />
=Works Cited=<br />
<br />
1. Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Turcotte, R., Raines, R., Interactions of Onconase with Human Ribonuclease Inhibitor. ''Biochemical Biophysical Research Communities'':(2008) Vol. 377, Iss. 4, pp. 512-414<br />
<br />
3. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''. ''Biochemistry'': (1996) Vol. 29, pp.8827-8834<br />
<br />
4. Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230526Ribonuclease A Catalysis2011-04-15T01:27:04Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/3'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/2'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/3'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''.<br />
<br />
3.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230521Ribonuclease A Catalysis2011-04-15T01:20:53Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the <scene name='Sandbox_Reserved_193/Active_site_a/1'>active site</scene> which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/1'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/1'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Arg10_a/1'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''.<br />
<br />
3.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230469Ribonuclease A Catalysis2011-04-15T00:08:47Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/1'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/1'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Arg10_a/1'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''.<br />
<br />
3.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230467Ribonuclease A Catalysis2011-04-15T00:06:36Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/1'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/1'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Arg10_a/1'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2. Vicentini,A., Protein Chemical and Kinetic Characterization of Recombinant Porcine Ribonuclease Inhibitor Expressed in ''Saccharomyces cerevisae''.<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230460Ribonuclease A Catalysis2011-04-14T23:51:56Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/1'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/1'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Arg10_a/1'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230459Ribonuclease A Catalysis2011-04-14T23:51:07Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/2 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/1'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/1'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Arg10_a/1'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
http://en.wikipedia.org/wiki/Ribonuclease_inhibitor<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230380Ribonuclease A Catalysis2011-04-14T19:20:33Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/Rnasei_a/1 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as <scene name='Sandbox_Reserved_193/Lys7_a/1'>Lys7</scene> and <scene name='Sandbox_Reserved_193/Lys66_a/1'>Lys66</scene> and <scene name='Sandbox_Reserved_193/Arg10_a/1'>Arg10</scene>, recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. <scene name='Sandbox_Reserved_193/Thr45_a/1'>Thr45</scene>, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
RNase A catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12a_a/1'>His12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41a_a/1'>Lys41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119a_a/1'>His119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230372Ribonuclease A Catalysis2011-04-14T18:55:47Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/His12/1 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230370Ribonuclease A Catalysis2011-04-14T18:55:11Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/His12/1 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230369Ribonuclease A Catalysis2011-04-14T18:54:43Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAIII.PNG|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/His12/1 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=File:RNaseAIII.png&diff=1230368File:RNaseAIII.png2011-04-14T18:54:17Z<p>Nathan Clarke: </p>
<hr />
<div></div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1230239Ribonuclease A Catalysis2011-04-14T00:29:31Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAII.PNG|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= Sandbox_Reserved_193/His12/1 }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
http://en.wikipedia.org/wiki/RNase_A<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1227516Ribonuclease A Catalysis2011-04-06T17:26:43Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAII.PNG|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechIII.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=File:MechIII.png&diff=1227515File:MechIII.png2011-04-06T17:26:09Z<p>Nathan Clarke: </p>
<hr />
<div></div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1227514Ribonuclease A Catalysis2011-04-06T17:22:50Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAII.PNG|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:MechII.PNG|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1227511Ribonuclease A Catalysis2011-04-06T17:00:49Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAII.PNG|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1227510Ribonuclease A Catalysis2011-04-06T16:59:10Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAII.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1227508Ribonuclease A Catalysis2011-04-06T16:52:29Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAI.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1227507Ribonuclease A Catalysis2011-04-06T16:51:47Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAI.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
<br />
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. Ribonuclease A, (RNase A), also uses acid/base catatalysis to chemically change its substrates. Acidic or basic residues of the enzyme transfer protons to or from the reactant in order to stabilize the developing charges in the transition state. The transfer of protons usually creates better leaving groups, making the reaction more energetically favorable. Histidine is a very common amino acid residue involved in cataylsis, as histidine has a pKa value close to neutral, (pKa=6); therefore, histidine can both accept and donate protons at physiological pH. <br />
<br />
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the pKa's of their residues. The pKa value will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
RNase A uses acid/base catlysis to speed up RNA hydrolysis. This occurs in the active site which is found in the cleft of RNase A and is the location of the chemical change in bound substrates. Subsites lining the active site cleft are important to the binding of single stranded RNA. Large quantities of positively charged residues, such as '''Lys7''' and '''Lys66''' and '''Arg10''', recognize the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A, although fairly nonspecific, has some specificity for sites RNA hydrolysis. '''Threonine 45''', located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. Thr45 significantly decreases the rate of hydrolysis of polymeric purine strands, such as poly A, by a thousand fold, as compared to polymeric pyrimidine strands. <br />
<br />
Early studies on RNase A catalysis showed that alkylation of His12 and His119 significantly decreased its catalytic activity, prompting the hypothesis that these two histidines were the acid/base catalyst. Confirmation of this hypothesis came when these histidines were replaced with alanine and the reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
<scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the Phosphodiester bonds in two steps: the formation of the pentavalent phosphate transition state and subsequent degradation of the 2’3’ cyclic phosphate intermediate. An important part of the reaction is the ability of histidine (His 12 and His119) to both accept and donate electrons, allowing these histidine to be an acid or a base, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
RNA hydrolysis begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> abstracts a proton from the 2’ OH group on RNA; thus, assisting in the nucleophilic attack of the 2’ oxygen on the electrophilic phosphorus atom. A transition state is then formed, having a pentavalent phosphate, which is stabilized by the positively charged amino group of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain amide nitrogen of Phe120. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> then protonates the 5' oxygen on the ribose ring and the transition state falls to form a 2’3’cyclic phosphate intermediate. <br />
<br />
<br />
In a secondary and separate reaction, the 2’,3’ cyclic phosphate is hydrolyzed to a mixture of 2'phosphate and 3' hydroxyl. His12 donates a proton to the leaving group of this reaction, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 abstracts the proton from a water molecule, activating it for nucleophilic attack. The activated water molecule attacks the cyclic phosphate causing the cleavage of the 2'3’ cyclic phosphate intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223717Ribonuclease A Catalysis2011-04-01T14:51:40Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAI.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. It is also a common mechanism by which enzyme cause chemical changes in their substrates. The acidic or basic residues are not consumed in the reaction themselves, but there is a transfer of protons to or from the reactant depending on the nature of the residue. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral, pKa 6; therefore, it can both accept and donate protons at physiological pH. <br />
<br />
The acid/base mechanism of enzymes are extensively dependent on the pH of the environment and the pKa's of their residues. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
The catalysis of RNA strands occurs in the active site which is the location of the chemical change in bound substrates. Subsites both up and down stream of the residues found around the active site are important for the binding of single stranded RNA. Large quantities of positively charged residues, such as Lys7 and 66 and Arg10, attract the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A actually has some specificity as to where it will degrade RNA stands. Threonine 45, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. This decreases the rates of polymeric purine strands, such as poly A, by a thousand fold.<br />
<br />
Early Studies on RNase A showed that alkylation of His12 and 119 showed marked decrease in the catalytic activity, prompting the notion that these were the active residues in catalysis. Through mutating these residues from Histidine to the chemically unreactive alanine, reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
In the acid base catalysis of RNA in mammalians, <scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the pentavalent phosphate transition state and subsequent degradation 2’3’ cyclic intermediate into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
The catalysis of RNase A begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> undergoes basic catalysis. His12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain of Phe120. This leads to the formation a stabilized 2’3’-cyclic intermediate. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degradation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223715Ribonuclease A Catalysis2011-04-01T14:46:35Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
[[Image:RNaseAI.png|300px|left|thumb|Figure I: Bovine Ribonuclease A. Colored residues are representative of amino acids important to both the acid base catalysis (Red: His12 and 119) and stabilization of the transition state (Blue: Lys41 and Phe120). Figure generated via ''Pymol'' ]]<br />
<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. It is also a common mechanism by which enzyme cause chemical changes in their substrates. The acidic or basic residues are not consumed in the reaction themselves, but there is a transfer of protons to or from the reactant depending on the nature of the residue. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral, pKa 6; therefore, it can both accept and donate protons at physiological pH. <br />
<br />
The acid/base mechanism of enzymes are extensively dependent on the pH of the environment and the pKa's of their residues. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. Having the ability to alter the pKa of certain residues such as Histidine, increases the diversity of reactions that can take place '''(reword)'''.<br />
<br />
<br />
===Active Site Structure===<br />
The catalysis of RNA strands occurs in the active site which is the location of the chemical change in bound substrates. Subsites both up and down stream of the residues found around the active site are important for the binding of single stranded RNA. Large quantities of positively charged residues, such as Lys7 and 66 and Arg10, attract the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
The active site for RNase A actually has some specificity as to where it will degrade RNA stands. Threonine 45, located next to the active site, will hydrogen bond to pyrimidine bases, but sterically hinder the binding of a purine on the 5' strand of OH. This decreases the rates of polymeric purine strands, such as poly A, by a thousand fold.<br />
<br />
Early Studies on RNase A showed that alkylation of His12 and 119 showed marked decrease in the catalytic activity, prompting the notion that these were the active residues in catalysis. Through mutating these residues from Histidine to the chemically unreactive alanine, reaction rates of either mutation dropped by ten-thousand fold. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
===Mechanism===<br />
In the acid base catalysis of RNA in mammalians, <scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the pentavalent phosphate transition state and subsequent degradation 2’3’ cyclic intermediate into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|450px|left|thumb|Figure II: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure generated via ''Chemdraw'']]<br />
<br />
<br />
The catalysis of RNase A begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> undergoes basic catalysis. His12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain of Phe120. This leads to the formation a stabilized 2’3’-cyclic intermediate. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
[[Image:Pentavalent.png|300px|right|thumb|Figure III: Pentavalent Phosphorous Transition State. Pictured borrowed from work by R. Raines]]<br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degradation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
===Inhibitors===<br />
Due to the highly catalytic nature of RNase A for RNA strands, mammalian cells have developed a protective inhibitor to prevent pancreatic ribonucleases from degrading cystolic RNA. Ribonuclease Inhibitor (RI) tightly associates to the active site due to its non-globular nature. '''I am waiting for a paper from nature that is pertinant to RIs. If any have to due with inhibition at the active site I thought this would be a good place to add. This is what I could get from the abstract.'''<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223500Ribonuclease A Catalysis2011-04-01T02:58:44Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
The catalysis of RNA strands occurs in the active site which is the location of the chemical change in bound substrates. Subsites both up and down stream of the residues found around the active site are important for the binding of single stranded RNA. Large quantities of positively charged residues, such as Lys 7 and 66 and Arg 10, attract the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
Substrate specificity of residues for RNA bases. <br />
<br />
HIS 12 and 119 and Lys 41.<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
<br />
In the acid base catalysis of RNA in mammalians, <scene name='Sandbox_Reserved_193/Rnase_a/3'>RNase A</scene> catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the pentavalent phosphate transition state and subsequent degradation 2’3’ cyclic intermediate into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|450px|left|thumb|Figure I: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His 12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences. Figure formed via ''Chemdraw'']]<br />
<br />
<br />
The catalysis of RNase A begins when <scene name='Sandbox_Reserved_193/His12/1'>His 12</scene> undergoes basic catalysis. H-12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of <scene name='Sandbox_Reserved_193/Lys41/1'> Lys 41</scene> and the main chain of Phe 120. This leads to the formation a stabilized 2’3’-cyclic intermediate. <scene name='Sandbox_Reserved_193/His119/1'>His 119</scene> will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
[[Image:Pentavalent.png|300px|right|thumb|Figure II: Pentavalent Phosphorous Transition State]]<br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His-12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His 119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degredation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223487Ribonuclease A Catalysis2011-04-01T02:33:47Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
The catalysis of RNA strands occurs in the active site which is the location of the chemical change in bound substrates. Subsites both up and down stream of the residues found around the active site are important for the binding of single stranded RNA. Large quantities of positively charged residues, such as Lys 7 and 66 and Arg 10, attract the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
<br />
In the acid base catalysis of RNA in mammalians, RNase A catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the pentavalent phosphate transition state and subsequent degradation 2’3’ cyclic intermediate into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|450px|left|thumb|Figure I: RNase A Catalysis. (A) Initial attack of 2'hydroxyl stabilized by His 12. (B) Pentavalent phosphorous intermediate. (C) 2'3' cyclic intermediate degradation. (D) Finished products: Two distinctive nucleotide sequences]]<br />
<br />
<br />
The catalysis of RNase A begins when His-12 undergoes basic catalysis. H-12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of Lys41 and the main chain of Phe120. This leads to the formation a stabilized 2’3’-cyclic intermediate. His-119 will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
[[Image:Pentavalent.png|300px|right|thumb|Figure II: Pentavalent Phosphorous Transition State]]<br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His-12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His 119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degredation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223475Ribonuclease A Catalysis2011-04-01T02:23:31Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
The catalysis of RNA strands occurs in the active site which is the location of the chemical change in bound substrates. Subsites both up and down stream of the residues found around the active site are important for the binding of single stranded RNA. Large quantities of positively charged residues, such as Lys 7 and 66 and Arg 10, attract the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
<br />
In the acid base catalysis of RNA in mammalians, RNase A catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the pentavalent phosphate transition state and subsequent degradation 2’3’ cyclic intermediate into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|500px|left|thumb|Figure I: RNase A Catalysis]]<br />
<br />
<br />
The catalysis of RNase A begins when His-12 undergoes basic catalysis. H-12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of Lys41 and the main chain of Phe120. This leads to the formation a stabilized 2’3’-cyclic intermediate. His-119 will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His-12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His 119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degredation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223465Ribonuclease A Catalysis2011-04-01T02:18:11Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
The catalysis of RNA strands occurs in the active site which is the location of the chemical change in bound substrates. Subsites both up and down stream of the residues found around the active site are important for the binding of single stranded RNA. Large quanities of positively charged residues, such as Lys 7 and 66 and Arg 10, attract the negative charge on the phosphate back bone of the RNA strand. <br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
<br />
In the acid base catalysis of RNA in mammalians, RNase A catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the 2’3’ cyclic intermediate and subsequent degradation of the RNA strand into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|500px|left|thumb|Figure I: RNase A Catalysis]]<br />
<br />
<br />
The catalysis of RNase A begins when His-12 undergoes basic catalysis. H-12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of Lys41 and the main chain of Phe120. This leads to the formation a stabilized 2’3’-cyclic intermediate. His-119 will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His-12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His 119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degredation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223433Ribonuclease A Catalysis2011-04-01T01:59:28Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
<br />
In the acid base catalysis of RNA in mammalians, RNase A catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the 2’3’ cyclic intermediate and subsequent degradation of the RNA strand into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|500px|left|thumb|Figure I: RNase A Catalysis]]<br />
<br />
<br />
The catalysis of RNase A begins when His-12 undergoes basic catalysis. H-12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of Lys41 and the main chain of Phe120. This leads to the formation a stabilized 2’3’-cyclic intermediate. His-119 will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His-12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His 119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degredation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
<br />
<br />
<br />
<br />
<br />
=Related Sites=<br />
<br />
=Works Cited=<br />
<br />
1.Raines, R. Ribonuclease A. ''Chemistry Review'': (1998) Vol. 98 pp. 1045-1068<br />
<br />
2.Wlodrawer, A., Svensson, L., Sjohin, L., Gilliland, G. Structure of Phosphate-Free Ribonuclease A Refined at 1.26A. ''Biochemistry'':(1988) Vol. 27 pp. 2705-2717</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223376Ribonuclease A Catalysis2011-04-01T00:57:00Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
===Acid Base Catalysis===<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
<br />
In the acid base catalysis of RNA in mammalians, RNase A catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the 2’3’ cyclic intermediate and subsequent degradation of the RNA strand into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|500px|left|thumb|Figure I: RNase A Catalysis]]<br />
<br />
<br />
The catalysis of RNase A begins when His-12 undergoes basic catalysis. H-12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of Lys41 and the main chain of Phe120. This leads to the formation a stabilized 2’3’-cyclic intermediate. His-119 will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His-12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His 119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degredation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223374Ribonuclease A Catalysis2011-04-01T00:54:16Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
===Acid Base Catalysis===<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
<br />
=='''Acid Base Catalysis by RNase A'''==<br />
<br />
In the acid base catalysis of RNA in mammalians, RNase A catalyzes the cleavage of the P-O 5’ bond, and is comprised of two separate processes, the formation of the 2’3’ cyclic intermediate and subsequent degradation of the RNA strand into its individual nucleotides. An important part of the reaction is Histidine’s ability to both accept and donate electrons. This acts as a proton source, allowing Histidine to be utilized as a base or acid, making the reaction pH dependent.<br />
<br />
<br />
[[Image:mech3.png|500px|left|thumb|Figure I: RNase A Catalysis]]<br />
<br />
<br />
The catalysis of RNase A begins when His-12 undergoes basic catalysis. H-12 will act as a base and abstract a proton from the RNA’s 2’ OH group; thus, assisting the attack of the 2’ oxygen on the phosphorus atom. This reaction occurs via a transition state, having a pentavalent phosphorous atom. These transition states are both stabilized by the side positive character of the side chain of Lys41 and the main chain of Phe120. This leads to the formation a stabilized 2’3’-cyclic intermediate. His-119 will support this reaction by protonating the leaving group, the 6’ OH on the ribose of the 3’ RNA, thus acting as a general acid. <br />
<br />
The 2’,3’- cyclic nucleotide is hydrolyzed in a separate process. His-12 will donate the excess proton from the initial step to the leaving group, the 3’ oxygen of the cyclic intermediate. Simultaneously, His-119 draws the hydrogen off of a water molecule. His 119 is thus reprotonated during this process, making water a better nucleophile. The water molecule attacks the phosphate causing the cleavage of the 2-3’ cyclic intermediate. The truncated nucleotide is then released with a 3’ phosphate group.<br />
Upon degredation of the phosphodiester linkage between the two nucleotides the products are then released into the surrounding solvent.<br />
<br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
<br />
=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223068Ribonuclease A Catalysis2011-03-31T16:23:40Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
===Acid Base Catalysis===<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223067Ribonuclease A Catalysis2011-03-31T16:22:47Z<p>Nathan Clarke: </p>
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{{Template:Johnson_CH462_Spring2011}}<br />
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<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
===Acid Base Catalysis===<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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<Structure load='7RSA' size='250' frame='' align='left' caption='Ribonuclease A' scene='Insert optional scene name here'/><br />
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=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223064Ribonuclease A Catalysis2011-03-31T16:19:44Z<p>Nathan Clarke: </p>
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{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
===Acid Base Catalysis===<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
===Structure===<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here'/><br />
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=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223063Ribonuclease A Catalysis2011-03-31T16:19:08Z<p>Nathan Clarke: </p>
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{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
===Acid Base Catalysis===<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
=Structure=<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here'/><br />
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=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223062Ribonuclease A Catalysis2011-03-31T16:18:37Z<p>Nathan Clarke: </p>
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{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
|Acid Base Catalysis|<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
=Structure=<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here'/><br />
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=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223061Ribonuclease A Catalysis2011-03-31T16:18:18Z<p>Nathan Clarke: </p>
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{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
'Acid Base Catalysis'<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
=Structure=<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here'/><br />
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=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223060Ribonuclease A Catalysis2011-03-31T16:17:15Z<p>Nathan Clarke: </p>
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<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
<br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
=Structure=<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here'/><br />
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=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223059Ribonuclease A Catalysis2011-03-31T16:13:44Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_7rsa | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here'/><br />
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<br />
<br />
<br />
<br />
=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223057Ribonuclease A Catalysis2011-03-31T16:12:35Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_3cin | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
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Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
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<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
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<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here'/><br />
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=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223056Ribonuclease A Catalysis2011-03-31T16:11:48Z<p>Nathan Clarke: </p>
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{{Template:Johnson_CH462_Spring2011}}<br />
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=Ribonuclease A Catalysis=<br />
==Introduction==<br />
{{STRUCTURE_3cin | PDB=7RSA | SCENE= }}<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
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The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
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==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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[[<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here'/>]]<br />
<br />
=References=<br />
=Works Cited=</div>Nathan Clarkehttps://proteopedia.org/index.php?title=Ribonuclease_A_Catalysis&diff=1223053Ribonuclease A Catalysis2011-03-31T16:10:04Z<p>Nathan Clarke: </p>
<hr />
<div><!-- PLEASE DO NOT DELETE THIS TEMPLATE --><br />
{{Template:Johnson_CH462_Spring2011}}<br />
<!-- PLEASE ADD YOUR CONTENT BELOW HERE --><br />
=Ribonuclease A Catalysis=<br />
<br />
<br />
{{STRUCTURE_3cin | PDB=7RSA | SCENE= }}<br />
<br />
==Introduction==<br />
Acid Base Catalysis is the acceleration of a chemical reaction by the addition of an acid or a base and is mainly used in organic chemical reactions. The acid or base is not consumed in the reaction itself. An acid transfers protons to a reactant and a base accepts protons from the reactant. An acid is often thought of as a proton and the base as a hydroxyl. When the acids or bases donate or accept protons, they stabilize the developing charges in the transition state. This usually creates a better leaving group, making the reaction more energetically favorable. Additionally, this has an effect on the activity of the nucleophile and electrophile groups. Histidine is a very common residue involved in acid-base cataylsis due to the fact that is has a pKa close to neutral; therefore, it can both accept and donate protons. <br />
<br />
The acid base mechanism can extensively alter the pKa depending on the environment of the residue. PKa will increase for an acidic residue if the environment is hydrophobic or if the adjacent residues are of similar charges. In the same environmental conditions, a basic residue will decrease the pKa. PKa will decrease for an acidic residue and increase for a basic residue if there is a salt bridge. <br />
<br />
<br />
==Mechanism==<br />
[[Image:mech3.png|400px|left|thumb|Figure I: Mechanism of RNase A Catalysis of Single Stranded RNA]] <br />
<br />
The catalysis of RNase A begins when Histidine 12 abstract a proton from the RNA’s 2’ OH group. (Talk about pKAs) This leads to the formation of the 2’3’-cyclic intermediate. In the active site of RNase A, Lysine 41 and Phenylalanine 120 stabilize the excess negative charge on the transition state. His-119 will support this reaction by protonating the leaving group: the 6’ OH on the ribose of the 3’ RNA strand. There is an excess positive charge on the imidazole ring of His-119, that stems from an extra hydrogen on N???. This hydrogen serves as the proton source for the leaving group.<br />
<br />
Hydrolysis of the 2’ 3’- cyclic intermediate occurs when the His-119 takes a proton from the surrounding water, which attacks the phosphorous of the phosphate group. Simultaneously His-12 supports this reaction by donating the extra proton from its imidazole ring, formed during the initial step, to the 2’ oxygen. Following the degradation of the complex, the two nucleotide strands are subsequently released from the active site. <br />
<br />
<br />
<scene name='Sandbox_Reserved_193/Rnase_a/2'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_119/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/His_12/1'>TextToBeDisplayed</scene><br />
<scene name='Sandbox_Reserved_193/Lys_41/1'>TextToBeDisplayed</scene><br />
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[[Image:<Structure load='7RSA' size='250' frame='true' align='right' caption='Ribonuclease A' scene='Insert optional scene name here' />]]<br />
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=References=<br />
=Works Cited=</div>Nathan Clarke