Using Corynebacterium Glutamicum To Efficiently Express Glutamine Synthetase And Fermentatively Produce L-glutamine

Corynebacterium glutamicum(C.glutamicum),the Gram-positive bacterium,one of the most important organisms in biotechnology,is usually used to produce amino acids L-lysine,L-glutamine(L-Gln),L-glutamate and sodium glutamate.Glutamine synthetase(GS)is the key enzyme responsible for L-glutamine(L-Gln)synthesis.However,enzymatic activity of GS is influenced by many factors,leading to the low enzyme activity and the low production of L-Gln.In this study,in order to improve the yield of L-Gln,a high-effective expression system was constructed in C.glutamicum with glnA gene.We tested two different strong promoters,promoter tac(Ptac),and promoter mal(Pmal),for a comparison of the activity of GS.After then,for avoiding the adenylylation in C.glutamicum,we constructed plasmids containing the point mutation of glnA,in which Tyr405 is replaced by a phenylalanine residue.Since the glutamine synthetase in Bacillus subtilis is not regulated via adenylylation,we also compared C.glutamicum GS activity with Bacillus subtilis’s to detect the enzyme activity in the given system.The results reveal that promoter tac was more capable for producing L-Gln in C.glutamicum system and GS enzyme from Bacillus subtilis has the lower activity than the C.glutamicum’s mutant.The recombinant strain BJ2 has the highest GS activity and produce 32.5 g/L L-Gln.