Effect of Tween 40 and DtsR1 on L-arginine overproduction in Corynebacterium crenatum.
BACKGROUND
L-Glutamate is an important precursor in the L-arginine (L-Arg) biosynthetic pathway. Various methods, including polyoxyethylene sorbitan monopalmitate (Tween 40) addition and dtsR1 disruption, have been widely used to induce L-glutamate overproduction in Corynebacterium glutamicum. In this study, a novel strategy for L-Arg overproduction through Tween 40 trigger and ΔdtsR1 mutant were proposed in Corynebacterium crenatum.


RESULTS
Corynebacterium crenatum mutant (CCM01) was selected as a host strain, whose argR was lethal via mutagenesis screening, the proB gene was knocked out, and argB was replaced by argB M4 (E19R, H26E, D311R, and D312R) to release L-Arg feedback resistance. After Tween 40 trigger in the logarithmic period, L-Arg production increased from 15.22 to 17.73 g/L in CCM01 strain. When NCgl1221 and dtsR1 disruption (CCM03), L-Arg production drastically increased to 27.45 g/L and then further to 29.97 g/L after Tween 40 trigger. Moreover, the specific activity of α-oxoglutarate dehydrogenase complex (ODHC) decreased, whereas the regeneration of NADP(+)/NADPH significantly increased after dtsR1 disruption and Tween 40 trigger. Results of real-time PCR showed that the transcriptional levels of odhA, sucB, and lpdA (encoding three subunits of the ODHC complex) were downregulated after Tween 40 trigger or dtsR1 disruption. By contrast, zwf transcription (encoding glucose-6-phosphate dehydrogenase) showed no significant difference among CCM01, CCM02 (ΔNCgl1221), and CCM03 (ΔNCgl1221ΔdtsR1) strains without Tween 40 trigger but evidently increased by 5.50 folds after Tween 40 trigger.


CONCLUSIONS
A novel strategy for L-Arg overproduction by dtsR1 disruption and Tween 40 trigger in C. crenatum was reported. Tween 40 addition exhibited a bifunctional mechanism for L-Arg overproduction, including reduced ODHC activity and enhanced NADPH pools accumulation by downregulated dtsR1 expression and upregulated zwf expression, respectively.
College of Life Science, Nanchang University, Nanchang, 330031, People's Republic of China. 13576094194@163.com.|State Key Laboratory of Food Science and Technology, Jiangxi-OAI Joint Research Institute, Nanchang University, 235 Nanjing East Road, Nanchang, 330047, People's Republic of China. 13576094194@163.com.
Minliang
M
Chen
Key Laboratory of Functional Small Organic Molecule, Ministry of Education, Jiangxi Normal University, 99 Ziyang Road, Nanchang, 330022, People's Republic of China. xuelanchen162@163.com.
Xuelan
X
Chen
Key Laboratory of Functional Small Organic Molecule, Ministry of Education, Jiangxi Normal University, 99 Ziyang Road, Nanchang, 330022, People's Republic of China. fcwy1314@163.com.
Fang
F
Wan
Key Laboratory of Functional Small Organic Molecule, Ministry of Education, Jiangxi Normal University, 99 Ziyang Road, Nanchang, 330022, People's Republic of China. zhangbin2912@163.com.
Bin
B
Zhang
College of Life Science, Nanchang University, Nanchang, 330031, People's Republic of China. m18070503204@163.com.|State Key Laboratory of Food Science and Technology, Jiangxi-OAI Joint Research Institute, Nanchang University, 235 Nanjing East Road, Nanchang, 330047, People's Republic of China. m18070503204@163.com.
Jincong
J
Chen
College of Life Science, Nanchang University, Nanchang, 330031, People's Republic of China. yhxiongchen@163.com.|State Key Laboratory of Food Science and Technology, Jiangxi-OAI Joint Research Institute, Nanchang University, 235 Nanjing East Road, Nanchang, 330047, People's Republic of China. yhxiongchen@163.com.
Yonghua
Y
Xiong
D001120:Arginine / Q000096:biosynthesis*; D003352:Corynebacterium / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D055786:Gene Knockout Techniques; D005798:Genes, Bacterial; D018698:Glutamic Acid / Q000096:biosynthesis*; D011136:Polysorbates / Q000494:pharmacology
D011136:Polysorbates; D018698:Glutamic Acid; D001120:Arginine; C068430:polysorbate 40