Formation and Stability of a Vinyl Carbanion at the Active Site of Orotidine 5‘-Monophosphate Decarboxylase: pKa of the C-6 Proton of Enzyme-Bound UMP
- 11 January 2008
- journal article
- research article
- Published by American Chemical Society (ACS) in Journal of the American Chemical Society
- Vol. 130 (5) , 1574-1575
- https://doi.org/10.1021/ja710384t
Abstract
We report that orotidine 5‘-monophosphate decarboxylase (OMPDC) catalyzes exchange of the C-6 proton of uridine 5‘-monophosphate (UMP) for deuterium from solvent in D2O at 25 °C and pD 7.0−9.3. Kinetic analysis of deuterium exchange gives pKa ≤ 22 for carbon deprotonation of enzyme-bound UMP, which is at least 10 units lower than that for deprotonation of an analogue of UMP in water. The observation of enzyme-catalyzed deuterium exchange via a stabilized carbanion provides convincing evidence for the decarboxylation of orotidine 5‘-monophosphate (OMP) by OMPDC to give the same carbanion intermediate. The data show that yeast OMPDC stabilizes the bound vinyl carbanion by at least 14 kcal/mol. We conclude that OMPDC also provides substantial stabilization of the late carbanion-like transition state for the decarboxylation of OMP, and that this transition state stabilization constitutes a large fraction, but probably not all, of the enormous 1017-fold enzymatic rate acceleration.Keywords
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