Microbial production of L -glutamate and L -glutamine by recombinant Corynebacterium glutamicum harboring Vitreoscilla hemoglobin gene vgb.
Vitreoscilla hemoglobin (VHb) gene vgb equipped with a native promoter Pvgb or a tac promoter Ptac was introduced into Corynebacterium glutamicum ATCC14067, respectively. Ptac was proven to be more suitable for expressing VHb protein in higher concentration in both Escherichia coli and C. glutamicum strains compared with the native vgb promoter Pvgb. VHb-expressing C. glutamicum exhibited higher oxygen uptake rate and enhanced cell growth. Recombinant C. glutamicum harboring vgb gene equipped with Ptac promoter produced 23% more L -glutamate in shake-flask culture and grew to 30% more cell density and formed 22% more L -glutamate in fermentor studies compared with the wild-type strain. When a site-directed mutagenesis in which Tyr405 was replaced by a phenylalanine residue (Y405F) was performed on glutamine synthesis gene, recombinant C. glutamicum overexpressing the mutated gene glnA' was able to produce L: -glutamine effectively. Co-expression of vgb and glnA' genes in C. glutamicum produced 17 g/l L -glutamine in shake flask culture, approximately 30% more than that produced by the recombinant harboring only glnA' gene. In fermentor cultivation, the recombinant yielded 25% more cells and produced 40.5 g/l L -glutamine. In this study, it was clearly demonstrated that VHb significantly enhanced cell growth, L -glutamate, and L -glutamine production by recombinant C. glutamicum.
Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing, China.
Qian
Q
Liu
Jiao
J
Zhang
Xiao-Xing
XX
Wei
Shao-Ping
SP
Ouyang
Qiong
Q
Wu
Guo-Qiang
GQ
Chen
D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics*; D018533:Biomass; D019149:Bioreactors; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D005973:Glutamine / Q000378:metabolism*; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D054793:Truncated Hemoglobins / Q000096:biosynthesis* / Q000235:genetics*
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D054793:Truncated Hemoglobins; D005973:Glutamine; C082613:hemoglobin protein, Vitreoscilla; D018698:Glutamic Acid