AUTHOR=Wu Yan , Chu Wanying , Yang Jiayao , Xu Yudong , Shen Qi , Yang Haoning , Xu Fangxu , Liu Yefei , Lu Ping , Jiang Ke , Zhao Hongxin 

TITLE=Metabolic Engineering of Enterobacter aerogenes for Improved 2,3-Butanediol Production by Manipulating NADH Levels and Overexpressing the Small RNA RyhB

JOURNAL=Frontiers in Microbiology

VOLUME=12

YEAR=2021

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2021.754306

DOI=10.3389/fmicb.2021.754306

ISSN=1664-302X

ABSTRACT=<p>Biotechnological production of 2,3-butanediol (2,3-BD), a versatile platform bio-chemical and a potential biofuel, is limited due to by-product toxicity. In this study, we aimed to redirect the metabolic flux toward 2,3-BD in <italic>Enterobacter aerogenes</italic> (<italic>E. aerogenes</italic>) by increasing the intracellular NADH pool. Increasing the NADH/NAD<sup>+</sup> ratio by knocking out the NADH dehydrogenase genes (<italic>nuoC</italic>/<italic>nuoD</italic>) enhanced 2,3-BD production by up to 67% compared with wild-type <italic>E. aerogenes</italic>. When lactate dehydrogenase (<italic>ldh</italic>) was knocked out, the yield of 2,3-BD was increased by 71.2% compared to the wild type. Metabolic flux analysis revealed that upregulated expression of the sRNA RyhB led to a noteworthy shift in metabolism. The 2,3-BD titer of the best mutant Ea-2 was almost seven times higher than that of the parent strain in a 5-L fermenter. In this study, an effective metabolic engineering strategy for improved 2,3-BD production was implemented by increasing the NADH/NAD<sup>+</sup> ratio and blocking competing pathways.</p>