Studies on the Mechanism of Myrosinase

Myrosinase (EC 3.2.3.1) is the beta-thioglucosidase enzyme responsible for the hydrolysis of glucosinolates, a group of naturally occurring plant metabolites. The enzyme catalyzes the hydrolysis of these S-glucosides to give D-glucose and an aglycone fragment, which then rearranges to give sulfate and an isothiocyanate. As part of ongoing mechanistic studies on myrosinase, the ability of the enzyme to catalyze transglycosylation reactions has been examined. Enzyme activity and stability were both decreased in the presence of various organic solvents, including simple alcohols, but not sufficiently to prevent reaction taking place. However, in contrast to most other beta-glycosidases, myrosinase did not catalyze transglycosylation reactions either with the alcohols or other suitable glycosyl acceptors. Although a wide range of potential acceptors were investigated, none proved to be effective. Even when appropriately charged side chains were included in the acceptor molecule to mimic the sulfonic acid in the glucosinolate structure, transglycosylation did not take place. The putative enzyme-glycosyl intermediate therefore appears to be unavailable for reaction, possibly because D-glucose is the first product released from the enzyme. The transition state analogue, glucono-delta-lactone, a potent competitive inhibitor of beta-glucosidase, was found to be a poor noncompetitive inhibitor of myrosinase. Myrosinase is specifically activated by ascorbic acid, and it is proposed that the inhibitor is binding at this alternative site.