Utilization of excess NADH in 2,3‐butanediol‐deficient Klebsiella pneumoniae for 1,3‐propanediol production

• Published in: Journal of applied microbiology 2014-09, Vol.117 (3), p.690-698
• Main Authors: Cui, Y.L, Zhou, J.J, Gao, L.R, Zhu, C.Q, Jiang, X, Fu, S.L, Gong, H
• Format: Article
• Language: English
• Subjects: 1,3‐propanediol; 2,3‐butanediol; Alcohol Dehydrogenase - genetics; Alcohol Dehydrogenase - metabolism; Bacteriology; Biological and medical sciences; Butylene Glycols - metabolism; cell growth; Cellular biology; dhaT overexpression; Fermentation; Fundamental and applied biological sciences. Psychology; gene overexpression; glycerol; Glycerol - metabolism; Klebsiella pneumoniae; Klebsiella pneumoniae - enzymology; Klebsiella pneumoniae - genetics; Klebsiella pneumoniae - metabolism; lactate; Lactic Acid - metabolism; metabolites; Microbiology; mutants; Mutation; NAD (coenzyme); NAD - metabolism; NADH; Propylene Glycols - metabolism
• ISSN: 1364-5072; 1365-2672
• DOI: 10.1111/jam.12588

AIMS: To utilize excess NADH for 1,3‐propanediol production by 2,3‐butanediol‐deficient mutants, the effect of dhaT overexpression in two distinct 2,3‐butanediol‐deficient mutants was investigated. METHODS AND RESULTS: Two 2,3‐butanediol‐deficient mutants, KG1‐3 (blocking of the 2,3‐butanediol pathway only) and KG1‐5 (blocking of both of 2,3‐butanediol and lactate pathways) were constructed. Our results showed that although the intracellular redox balance (NADH/NAD⁺) was extremely high at the end of fermentation for both mutants, the status of intracellular redox in KG1‐5 was maintained at a normal level following the first stage of fermentation. Analysis of cell growth and metabolite formation confirmed the inhibition of excess lactate in 2,3‐butanediol pathway‐deficient mutants. Furthermore, dhaT was overexpressed in two 2,3‐butanediol‐deficient mutants (KG1‐3T and KG1‐5T). In KG1‐5T, the intracellular redox balance was restored to normal and 1,3‐propanediol production increased. The yield of 1,3‐propanediol from glycerol in KG1‐5T was also restored to a normal level of 0·6. CONCLUSIONS: The excess NADH in both the 2,3‐butanediol‐ and lactate‐deficient mutants can be used by overexpresstion of dhaT. SIGNIFICANCE AND IMPACT OF STUDY: The metabolic flux tended to increase lactate production by the abolishment of the 2,3‐butanediol pathway in Klebsiella pneumoniae, and the high accumulation of lactate prevented the cell from using excess NADH, thereby inhibiting cell growth and 1,3‐propanediol production.