Citryl Phosphate and the Mode of Action of the Citrate Cleavage Enzyme

When the citrate cleavage reaction is conducted with 18O-citrate, 1 atom of citrate oxygen is incorporated into each molecule of orthophosphate stemming from ATP. This is consistent with the notion that citryl phosphate is an intermediate of the reaction. Chemically synthesized dl-citryl 1-phosphate is an active substrate of the citrate cleavage enzyme, undergoing stereospecific cleavage in the presence of coenzyme A to oxalacetate and acetyl coenzyme A. Citryl phosphate transfers its citryl group to the enzyme to form citryl enzyme, which is identical in its biochemical properties with the citryl enzyme formed from citrate plus ATP. Citryl phosphate can also transfer its phosphoryl group to the enzyme to form a phosphoprotein which differs in its properties from the phosphoenzyme formed from ATP.