Studies on a possible phosphoryl-enzyme intermediate in the catalytic reaction of yeast phosphoglycerate kinase

A phosphoryl-enzyme intermediate as part of the mechanism of phosphoglycerate kinase has been suggested for the rabbit muscle enzyme (6) and the yeast enzyme (7,8). ATP in the binary enzyme-substrate complexes appeared to phosphorylate these enzymes and ADP-ATP exchange activities were observed (6,7,8). The present report shows, however, that highly purified yeast enzyme cannot be phosphorylated by ATP. On the other hand ADP-ATP exchange activity was obtained but this was proportional to trace amounts of adenylate kinase activity, which was found to contaminate the enzyme preparations. Thus a Ping Pong mechanism as an alternative to a mechanism including a ternary complex between the enzyme and its two substrates appears very improbable. Whether the enzyme or the phosphoryl-group-accepting substrate is responsible for the primary nucleophilic attack occurring in the ternary complex is still an open question, however. Yeast phosphoglycerate kinase appears to have no ATPase activity.