Evolutionary engineering of Escherichia coli for improved anaerobic growth in minimal medium accelerated lactate production
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Anaerobic fermentation is a favorable process for microbial production of bulk chemicals like ethanol and organic acids. Low productivity is the bottleneck of several anaerobic processes which has significant impact on the technique competitiveness of production strain. Improving growth rate of production strain can speed up the total production cycle and may finally increase productivity of anaerobic processes. In this work, evolutionary engineering of wild-type strain Escherichia coli W3110 was adopted to improve anaerobic growth in mineral medium. Significant increases in exponential growth rate and stationary cell density were achieved in evolved strain WE269, and a 96.5% increase in lactate productivity has also been observed in batch fermentation of this strain with M9 minimal medium. Then, an engineered strain for lactate production (BW100) was constructed by using WE269 as a platform and 98.3 g/L lactate (with an optical purity of D-lactate above 95%) was produced in a 5-L bioreactor after 48 h with a productivity of 2.05 g/(L·h). Finally, preliminary investigation demonstrated that mutation in sucD (sucD M245I) (encoding succinyl-CoA synthetase); ilvG (ilvG Δ1bp) (encoding acetolactate synthase 2 catalytic subunit), and rpoB (rpoB T1037P) (encoding RNA polymerase β subunit) significantly improved anaerobic growth of E. coli. Double-gene mutation in ilvG and sucD resumed most of the growth potential of evolved strain WE269. This work suggested that improving anaerobic growth of production host can increase productivity of organic acids like lactate, and specific mutation-enabled improved growth may also be applied to metabolic engineering for production of other bulk chemicals.
KeywordsEvolutionary engineering Escherichia coli Anaerobic growth Productivity sucD rpoB Lactate
We thank Prof. Qinghong Wang (TIB, CAS) for his kind gift of wild-type strain of this study. We also thank Dr. Zhubo Dai (TIB, CAS) and Yufeng Mao (TJU, China) for discussion on whole genome resequencing analysis.
This work was supported by the National Natural Science Foundation of China (NSFC-21776208, NSFC- 21621004, NSFC-21776209 and NSFC-21390201).
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interests.
This article does not contain any studies with human participants or animals performed by any of the authors.
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