RNase A: Difference between revisions
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=='''Ribonuclease A Catalysis'''== | =='''Ribonuclease A Catalysis'''== | ||
==='''Active Site Structure'''=== | ==='''Active Site Structure'''=== | ||
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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 = 'Wlodrawer'>PMID:3401445</ref> | 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 = 'Wlodrawer'>PMID:3401445</ref> | ||
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="Wlodrawer" />. | 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="Wlodrawer" />. | ||
==='''Acid Base Catalysis'''=== | |||
In organic chemistry acid/base catalysis is the addition of an acid or base to accelerate a chemical reaction. 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, (p''K''a=6); therefore, histidine can both accept and donate protons at physiological pH. | |||
Acid/base catalysis by an enzyme is dependent on the pH of the environment and the p''K''a'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 p''K''a. This ability to alter the pKa of certain residues such as histidines, increases the diversity of reactions that an enzyme can perform <ref name = 'Lehninger'>'Lehninger A., Nelson D.N, & Cox M.M. (2008) Lehninger Principles of Biochemistry. W. H. Freeman, fifth edition.' </ref>. | |||
==='''Acid Base Catalysis by RNase A'''=== | ==='''Acid Base Catalysis by RNase A'''=== | ||
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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="Raines" />. | 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="Raines" />. | ||
== '''Ribonuclease A Substrate Binding''' == | == '''Ribonuclease A Substrate Binding''' == | ||
