title: Regulation of the NADH pool and NADH/NADPH ratio redistributes acetoin and 2,3-butanediol proportion in Bacillus subtilis.
abstract: Bacillus subtilis produces acetoin as a major product along with several NADH-dependent byproducts, especially 2,3-butanediol. In this study, the down-regulation of the NADH pool and the redistribution of NADH/NADPH were targeted using external and genetic processes, as a means by which to redistribute the metabolic flux in favor of acetoin synthesis. First, it was found that the use of carbon sources of different oxidation states resulted in very different intracellular NADH/NAD(+) ratios that dictated the total process yield of acetoin. A mixture of glucose and gluconate as substrate produced a relatively low NADH/NAD(+) ratio, and resulted in an increase in acetoin production while byproducts significantly decreased. Metabolic engineering methods using glucose as a substrate could yield a similar effect. Acetoin production was significantly enhanced by overexpression of the oxidative pentose phosphate pathway: increased expression of glucose-6-phosphate dehydrogenase resulted in a decrease in the intracellular NADH/NADPH ratio (1.9-fold) and NADH/NAD(+) ratio (1.7-fold). In fed-batch culture the engineered strain yielded an acetoin concentration of 43.3 g L(-1) , while the production of 2,3-butanediol was only 1.7 g L(-1) . The concept of the manipulation of cofactor levels to redistribute carbon flux by external and genetic means as explored in this paper provides a novel strategy for improving industrial acetoin fermentation.
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu, China.
authors.forename: Teng
authors.initials: T
authors.lastname: Bao
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu, China.
authors.forename: Xian
authors.initials: X
authors.lastname: Zhang
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu, China.
authors.forename: Xiaojing
authors.initials: X
authors.lastname: Zhao
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu, China. raozhm@jiangnan.edu.cn.
authors.forename: Zhiming
authors.initials: Z
authors.lastname: Rao
authors.affiliation: The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu, China.
authors.forename: Taowei
authors.initials: T
authors.lastname: Yang
authors.affiliation: Department of Chemical Engineering, Ohio State University, Columbus, Ohio, USA.
authors.forename: Shangtian
authors.initials: S
authors.lastname: Yang
mesh_terms: D000093:Acetoin / Q000378:metabolism*; D001412:Bacillus subtilis / Q000235:genetics / Q000378:metabolism*; D001709:Biotechnology; D002072:Butylene Glycols / Q000378:metabolism*; D005285:Fermentation; D042843:Glucose 1-Dehydrogenase / Q000378:metabolism; D005954:Glucosephosphate Dehydrogenase / Q000378:metabolism; D060847:Metabolic Engineering; D009243:NAD / Q000378:metabolism*; D009249:NADP / Q000378:metabolism*
chemical_list: D002072:Butylene Glycols; D009243:NAD; C026978:2,3-butylene glycol; D009249:NADP; D000093:Acetoin; D042843:Glucose 1-Dehydrogenase; D005954:Glucosephosphate Dehydrogenase
keywords: Acetoin; Bacillus subtilis; Gluconate; Glucose-6-phosphate dehydrogenase; NADH/NAD+