A nucleophile activation dyad in ribonucleases. A combined X-ray crystallographic/ab initio quantum chemical study.
Ribonucleases (RNases) catalyze the cleavage of the phosphodiester bond in RNA up to 10(15)-fold, as compared with the uncatalyzed reaction. High resolution crystal structures of these enzymes in complex with 3'-mononucleotide substrates demonstrate the accommodation of the nucleophilic 2'-OH group in a binding pocket comprising the catalytic base (glutamate or histidine) and a charged hydrogen bond donor (lysine or histidine). Ab initio quantum chemical calculations performed on such Michaelis complexes of the mammalian RNase A (EC ) and the microbial RNase T(1) (EC ) show negative charge build up on the 2'-oxygen upon substrate binding. The increased nucleophilicity results from stronger hydrogen bonding to the catalytic base, which is mediated by a hydrogen bond from the charged donor. This hitherto unrecognized catalytic dyad in ribonucleases constitutes a general mechanism for nucleophile activation in both enzymic and RNA-catalyzed phosphoryl transfer reactions.
Mignon
Pierre
P
Eenheid Algemene Chemie (ALGC), Faculteit Wetenschappen, Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussels, Belgium.
Steyaert
Jan
J
Loris
Remy
R
Geerlings
Paul
P
Loverix
Stefan
S
D001665:Binding Sites; D018360:Crystallography, X-Ray / Q000379:methods*; D006860:Hydrogen Bonding; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008956:Models, Chemical; D008958:Models, Molecular; D010100:Oxygen / Q000378:metabolism; D011485:Protein Binding; D006163:Ribonuclease T1 / Q000737:chemistry*; D012259:Ribonuclease, Pancreatic / Q000737:chemistry*; D012260:Ribonucleases / Q000737:chemistry*; D013379:Substrate Specificity
D012260:Ribonucleases; D006163:Ribonuclease T1; D012259:Ribonuclease, Pancreatic; D010100:Oxygen