Overview of RNase A Structure/Function Relationships

Ribonuclease A (RNase A) is a small (124 amino acids) digestive enzyme secreted by the pancreas and is an endonuclease specifically designed to hydrolyze RNA (but not DNA) phosphodiester bonds which covalently link ribonucleotides particularly those linked to pyrimidine bases such as uracil.

Two His residues -- His12 and His119 -- are implicated in the catalytic mechanism of this enzyme. The imidazole ring of His12 has an anonymously low pK value (pk < 6.0) suggesting it must be deprotonated for catalysis. Conversely, the imidazole ring of His119 has an anonymously high pK value (pk > 6.0) suggesting it must be protonated for catalysis.

A two-step reaction is proposed for hydolysis of RNA.

1. His12 acts as a general base accepting 2'-OH proton of the sessile ribonucleic sugar ring thus promoting a nucleophilic attack by the 2'-O on the more positively-charged phosphorus (P) atom thereby creating a 2’,3’-cyclic ribonucleotide phosphate intermediate. The creation of this intermediate is facilitated by the imidazole of His119 which acts as a general acid donating its proton to the oxygen atom in the susceptible P-O-R' bond.
2. For the second step of the reaction, the general acid and base activities of the sidechains of these two His residue is reversed. His12 acts as a general acid donating its newly acquired proton to the 2’,3’-cyclic ribonucleotide phosphate intermediate while His119 which acts as a general base accepting a proton from water to promote hydroxyl attack on the same 2’,3’-cyclic intermediate

Uridine vanadate is a potent competitive inhibitor of RNase A. What makes it so potent is the fact that its structure mimicks a transition state intermediate in the reaction. Specifically, the oxidized vanadium atom is pentavalent ion with five coordinated oxygen atoms whose stable structure is believed to mimic the bipyramid structure of an unstable pentavalent phosphodiester intermediate that transiently forms in the active site of the enzyme.

Legend: 2',3'-cyclic uridine vanadate complex with RNase A active site. The pentacovalently coordinated vanadium (V) atom of the inhibitor forms metal bonds with 5 oxygen atoms (redspheres) creating a bipyramidal 2',3' cyclic ribonucleotide (black wireframe) linked to uracil. Specific active site residue contacts with His 119, His12, Lys41, and Thr45 are shown with distance measurements indicated in Ångstroms.

Additional information concerning the properties of RNase A and its interaction with another inhibitor (TBU), 2-methyl-2-proponol (t-butyl alcohol), can be also found at the following world-wide web sites:

• General properties of RNase A
• RNase A/TBU inhibitor complex

Go to the MCDB 108AL RNase A web page for further examination of the properties of this enzyme.

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©	Duane W. Sears Revised: July 27, 1998