Abstract
An enzyme preparation catalyzing p-nitroaniline release from γ-glutamyl-p-nitroanilide was obtained in a 200-fold purified state from fruit bodies of an edible mushroom, Lentinus edodes. Analysis of the final preparation by differential centrifugation revealed that the enzyme was still bound with subcellular particles. The enzyme catalyzed both the hydrolysis and transfer of the γ-glutamyl moiety from γ-glutamyl-p-nitroanilide, but exhibited essentially no activity of glutaminase, glutamine aminotransferase, glutamine synthetase or γ-glutamyl cyclotransferase. With γ-glutamyl-p-nitroanilide the activity was maximal at about pH 7.6. The enzyme activity increased with an increasing concentration of Tris-HCl buffer, but not with phosphate buffer which was inhibitory. An apparent Michaelis constant of 4mM was obtained in 0.5M Tris-HCl buffer at pH 7.6. S-Alkylcysteine sulfoxide served as the best glutamyl acceptor. A serine-borate mixture, pCMB, Cu2+, Hg2+ and Zn2+ were potent inhibitors. All the experimental results, including the insoluble nature of the enzyme, allowed us to classify the Lentinus enzyme in the family of γ-glutamyl transferase.
