pH-Dependent hydrolase, glutaminase, transpeptidase and autotranspeptidase activities of Bacillus subtilis γ-glutamyltransferase

-Glutamyltransferases (-GTs) are heterodimeric enzymes that catalyze the transfer of a -glutamyl group from a donor species to an acceptor molecule in a transpeptidation reaction through the formation of an intermediate -glutamyl enzyme. In our search for a -GT from a generally recognized as safe microorganism suitable for the production of -glutamyl derivatives with flavor-enhancing properties intended for human use, we cloned and overexpressed the -GT from Bacillussubtilis. In this study, we report the behavior of B.subtilis -GT in reactions involving glutamine as the donor compound and various acceptor amino acids. The common thread emerging from our results is a strong dependence of the hydrolase, transpeptidase and autotranspeptidase activities of B.subtilis -GT on pH, also in relation to the pK(a) of the acceptor amino acids. Glutamine, commonly referred to as a poor acceptor molecule, undergoes rapid autotranspeptidation at elevated pH, affording oligomeric species, in which up to four -glutamyl moieties are linked to a single glutamine. Moreover, we found that d-glutamine is also recognized both as a donor and as an acceptor substrate. Our results prove that the B.subtilis -GT-catalyzed transpeptidation reaction is feasible, and the observed activities of -GT from B.subtilis could be interpreted in relation to the known ability of the enzyme to process the polymeric material -polyglutamic acid.