Enhancement of glutathione production with a tripeptidase-deficient recombinant Escherichia coli.
Glutathione (GSH) degradation exists in the enzymatic synthesis of GSH by Escherichia coli, however, its degradation pathway is not very clear. This paper examines the key enzymes responding to GSH degradation in E. coli with the purpose of improving GSH production. The enzymes that are probably associated with GSH degradation were investigated by disrupting their genes. The results suggested that gamma-glutamyltranspeptidase (GGT) and tripeptidase (PepT) were the key enzymes of GSH degradation, and GGT contributed more to GSH degradation than PepT. Furthermore, GGT activity was affected greatly by culture temperature, and the effect of GGT on GSH degradation could be eliminated by shortening the culture time at 30 degrees C and extending the induction time at 42 degrees C. However, the effect of PepT on GSH degradation could be eliminated only by disrupting the PepT gene. Finally, GSH degradation was not observed in GSH biosynthesis by E. coli JW1113 (pepT(-), pBV03), which was cultured at 30 degrees C for 3 h and 42 degrees C for 5 h. GSH concentration reached 15.60 mM, which was 2.19-fold of the control. To the best of our knowledge, this is the first report of prohibiting GSH degradation with PepT-deficient recombinant E. coli. The results are helpful to investigate the GSH metabolism pathway and construct a GSH biosynthesis system.
Lin
Jun
J
Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, China.
Liao
Xianyan
X
Zhang
Juan
J
Du
Guocheng
G
Chen
Jian
J
D000626:Aminopeptidases / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D055786:Gene Knockout Techniques; D005978:Glutathione / Q000096:biosynthesis* / Q000378:metabolism; D008969:Molecular Sequence Data; D017510:Protein Folding; D005723:gamma-Glutamyltransferase / Q000235:genetics / Q000378:metabolism
D005723:gamma-Glutamyltransferase; D000626:Aminopeptidases; C023191:PepT tripeptidase; D005978:Glutathione