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

gamma-Glutamyltransferases (gamma-GTs) are heterodimeric enzymes that catalyze the transfer of a gamma-glutamyl group from a donor species to an acceptor molecule in a transpeptidation reaction through the formation of an intermediate gamma-glutamyl enzyme. In our search for a gamma-GT from a generally recognized as safe microorganism suitable for the production of gamma-glutamyl derivatives with flavor-enhancing properties intended for human use, we cloned and overexpressed the gamma-GT from Bacillus subtilis. In this study, we report the behavior of B. subtilis c-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 c-GT on pH, also in relation
to the pKa 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 gamma-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 gamma-GT-catalyzed transpeptidation reaction is feasible, and the observed activities of gamma-GT from B. subtilis could be interpreted in relation to the known ability of the enzyme to process the polymeric material gamma-polyglutamic acid.