Enhancing succinic acid biosynthesis in Escherichia coli by engineering its global transcription factor, catabolite repressor/activator (Cra).
This study was initiated to improve E. coli succinate production by engineering the E. coli global transcription factor, Cra (catabolite repressor/activator). Random mutagenesis libraries were generated through error-prone PCR of cra. After re-screening and mutation site integration, the best mutant strain was Tang1541, which provided a final succinate concentration of 79.8 ± 3.1 g/L: i.e., 22.8% greater than that obtained using an empty vector control. The genes and enzymes involved in phosphoenolpyruvate (PEP) carboxylation and the glyoxylate pathway were activated, either directly or indirectly, through the mutation of Cra. The parameters for interaction of Cra and DNA indicated that the Cra mutant was bound to aceBAK, thereby activating the genes involved in glyoxylate pathway and further improving succinate production even in the presence of its effector fructose-1,6-bisphosphate (FBP). It suggested that some of the negative effect of FBP on Cra might have been counteracted through the enhanced binding affinity of the Cra mutant for FBP or the change of Cra structure. This work provides useful information about understanding the transcriptional regulation of succinate biosynthesis.
School of Public Health, Wuhan University, Wuhan 430071 China.|Key Laboratory of Fermentation Engineering (Ministry of Education), Hubei Key Laboratory of Industrial Microbiology, Hubei Provincial Cooperative Innovation Center of Industrial Fermentation, Hubei University of Technology, Wuhan 430068 China.
Li-Wen
LW
Zhu
Key Laboratory of Fermentation Engineering (Ministry of Education), Hubei Key Laboratory of Industrial Microbiology, Hubei Provincial Cooperative Innovation Center of Industrial Fermentation, Hubei University of Technology, Wuhan 430068 China.
Shi-Tao
ST
Xia
Key Laboratory of Fermentation Engineering (Ministry of Education), Hubei Key Laboratory of Industrial Microbiology, Hubei Provincial Cooperative Innovation Center of Industrial Fermentation, Hubei University of Technology, Wuhan 430068 China.
Li-Na
LN
Wei
Key Laboratory of Fermentation Engineering (Ministry of Education), Hubei Key Laboratory of Industrial Microbiology, Hubei Provincial Cooperative Innovation Center of Industrial Fermentation, Hubei University of Technology, Wuhan 430068 China.
Hong-Mei
HM
Li
School of Public Health, Wuhan University, Wuhan 430071 China.
Zhan-Peng
ZP
Yuan
Key Laboratory of Fermentation Engineering (Ministry of Education), Hubei Key Laboratory of Industrial Microbiology, Hubei Provincial Cooperative Innovation Center of Industrial Fermentation, Hubei University of Technology, Wuhan 430068 China.
Ya-Jie
YJ
Tang
D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D005635:Fructosediphosphates / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D006038:Glyoxylates / Q000378:metabolism; D009154:Mutation / Q000502:physiology; D012097:Repressor Proteins / Q000378:metabolism*; D019802:Succinic Acid / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology
D029968:Escherichia coli Proteins; D005635:Fructosediphosphates; D006038:Glyoxylates; D012097:Repressor Proteins; D014157:Transcription Factors; D019802:Succinic Acid; C031150:glyoxylic acid; C029063:fructose-1,6-diphosphate