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 degrees 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.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Annotation Extension||Reference|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Assay||Construct||Conditions||Strain Background||Reference|