One-step fermentation converting glucose in γ-aminobutyric acid by a recombinant Corynebacterium crenatum

Abstract:

Abstract:［Objective］We constructed an integrative recombinant Corynebacterium crenatum that could directly convert glucose in γ-aminobutyric acid (GABA)．［Methods］ Using overlap PCR，we obtained ΔargB fragment that lacked 491 bp of N-acetylglutamate kinase (NAGK) gene． The glutamate decarboxylase (GAD) gene attached tac promoter was amplified and inserted into ΔargB fragment． Using the upstream and downstream of ΔargB as homologous arms，we constructed pK18-ΔargB:: tacgad which was used for the integration of tacgad gene onto the C．crenatum genome．Through two homologous recombinations，we got an argB blocked strain C．crenatum ΔargB:: tacgad which could successfully express GAD． We also fermented the recombinant strain with glucose as substrate and the production of GABA was detected．［Results］In C．crenatum ΔargB::tacgad，NAGK was totally inactivated and no L-arginine was detected though L-glutamic acid was accumulated. As a result of the efficient expression of GAD，part of L-glutamic acid was transformed into GABA，and the final yield of GABA was about 8. 28 g /L.［Conclusion］We successfully constructed a recombinant strain that could efficiently produce GABA by one-step fermentation from glucose． This research provided a new approach for GABA production.