First Example of the Extracellular Surface Expression of Intrinsically Periplasmic Escherichia coli γ-Glutamyltranspeptidase, a Member of the N-Terminal Nucleophile Hydrolase Superfamily, and the Use of Cells as a Catalyst for γ-Glutamylvalylglycine Production

Although the purified Escherichia coli gamma-glutamyltranspeptidase has much higher transpeptidation activity than hydrolysis activity, almost all gamma-glutamyltranspeptidase activity is hydrolysis activity in vivo, that is when measured using the whole cells. By using the Met1 to Arg232 fragment of E. coli YiaT or the CapA of Bacillus subtilis subsp. Natto as an anchor protein, we succeeded in expressing E. coli gamma-glutamyltranspeptidase on the extracellular surface of the cells, and these cells showed higher transpeptidation activity than hydrolysis activity in the presence of NaCl. Furthermore, E. coli cells overexpressing gamma- glutamyltranspeptidase without an anchor from the T5 promoter maintained gamma-glutamyltranspeptidase on the extracellular surface of the cells immediately after being harvested from the culture medium, but the enzyme was released from the extracellular surface of the cells subsequently in the absence of NaCl. Using these cells expressing gamma-glutamyltranspeptidase on the extracellular surface, gamma- Glu-Val-Gly, a kokumi compound, was successfully produced.

Automated summary

Although the purified Escherichia coli gamma-glutamyltranspeptidase exhibits higher transpeptidation activity than hydrolysis activity, most of the gamma-glutamyltranspeptidase activity in vivo is hydrolysis activity when measured using whole cells. By employing specific anchor proteins, E. coli cells were able to express gamma-glutamyltranspeptidase on the extracellular surface, resulting in higher transpeptidation activity compared to hydrolysis activity in the presence of NaCl. These cells were also used successfully to produce a kokumi compound, gamma-Glu-Val-Gly.