The mechanism of the reaction of cysteine with pyridoxal 5′-phosphate

Abstract

The reactions of pyridoxal 5′-phosphate with l-cysteine, S-methyl- l-cysteine, l-cysteine betaine and β-mercaptopropionic acid have been studied at various temperatures and at two concentrations of hydrogen ion. Methods have been developed for the determination, from optical data, of equilibrium constants for Schiff base, hemimercaptal, and mercaptal formation under suitable experimental conditions. It is shown that the forward and backward rate constants, k a and k b for Schiff base formation between pyridoxal 5′-phosphate and S-methyl- l-cysteine can be measured. The second-order rate constants, k a ′, for the cyclization of l-cysteine with pyridoxal 5′-phosphate, under the various conditions of temperature and pH, have also been determined. The values of k a and k a ′ are practically equal. No Schiff base was detectable in reaction mixtures of l-cysteine and pyridoxal 5′-phosphate. The equilibrium constants for hemimercaptal formation of pyridoxal 5′-phosphate with l-cysteine betaine and β-mercaptopropionic acid were small. These facts are strong evidence in favor of the hypothesis that the formation of a cyclic product between l-cysteine and pyridoxal 5′-phosphate occurs via the Schiff base, and that the formation of the latter is rate controlling.