Abstract
BackgroundWater-forming NADH oxidase can oxidize cytosolic NADH to NAD+, thus relieving cytosolic NADH accumulation in Saccharomyces cerevisiae. Previous studies of the enzyme were conducted under aerobic conditions, as O2 is the recognized electron acceptor of the enzyme. In order to extend its use in industrial production and to study its effect on anaerobes, the effects of overexpression of this oxidase in S. cerevisiae BY4741 and Clostridium acetobutylicum 428 (Cac-428) under anaerobic conditions were evaluated.ResultsGlucose was exhausted in the NADH oxidase-overexpressing S. cerevisiae strain (Sce-NOX) culture after 26 h, while 43.51 ± 2.18 g/L residual glucose was left in the control strain (Sce-CON) culture at this time point. After 30 h of fermentation, the concentration of ethanol produced by Sce-NOX reached 36.28 ± 1.81 g/L, an increase of 56.38 % as compared to Sce-CON (23.20 ± 1.16 g/L), while the byproduct glycerol was remarkably decreased in the culture of Sce-NOX. In the case of the C. acetobutylicum strain (Cac-NOX) overexpressing NADH oxidase, glucose consumption, cell growth rate, and the production of acetone–butanol–ethanol (ABE) all decreased, while the concentrations of acetic acid and butyric acid increased as compared to the control strain (Cac-CON). During fermentation of Cac-CON and Cac-NOX in 100-mL screw-capped bottles, the concentrations of ABE increased with increasing headspace. Additionally, several alternative electron acceptors in C. acetobutylicum fermentation were tested. Nitroblue tetrazolium and 2,6-dichloroindophenol were lethiferous to both Cac-CON and Cac-NOX. Methylene blue could relieve the effect caused by the overexpression of the NADH oxidase on the metabolic network of C. acetobutylicum strains, while cytochrome c aggravated the effect.ConclusionsThe water-forming NADH oxidase could regulate the metabolism of both the S. cerevisiae and the C. acetobutylicum strains in anaerobic conditions. Thus, the recombinant S. cerevisiae strain might be useful in industrial production. Besides the recognized electron acceptor O2, methylene blue and/or the structural analogs may be the alternative elector acceptor of the NADH oxidase in anaerobic conditions.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0517-y) contains supplementary material, which is available to authorized users.
Highlights
Water-forming NADH oxidase can oxidize cytosolic NADH to NAD+, relieving cytosolic NADH accumulation in Saccharomyces cerevisiae
Our results showed that overexpression of the NADH oxidase could regulate the metabolism of both the S. cerevisiae and the C. acetobutylicum strains in anaerobic condition, which can be generalized to other strains
Alternative electron acceptors of the water‐forming NADH oxidase in anaerobic condition Our results indicated that the water-forming NADH oxidase has alternative electron acceptors in anaerobic condition
Summary
Water-forming NADH oxidase can oxidize cytosolic NADH to NAD+, relieving cytosolic NADH accumulation in Saccharomyces cerevisiae. Previous studies of the enzyme were conducted under aerobic conditions, as O2 is the recognized electron acceptor of the enzyme. In order to extend its use in industrial production and to study its effect on anaerobes, the effects of overexpression of this oxidase in S. cerevisiae BY4741 and Clostridium acetobutylicum 428 (Cac-428) under anaerobic conditions were evaluated. Previous studies showed that overexpression of the water-forming NADH oxidase could increase the consumption of glucose and decrease the accumulation of glycerol in aerobic fermentation [1, 2]. All studies of this enzyme have been conducted under aerobic conditions, because O2 is recognized as the optimal electron acceptor of the enzyme. This study aimed to investigate whether the oxidase works under anaerobic conditions
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