Abstract
BackgroundRedox cofactors of NADH/NADPH participate in many cellular metabolic pathways for facilitating the electron transfer from one molecule to another in redox reactions. Transhydrogenase plays an important role in linking catabolism and anabolism, regulating the ratio of NADH/NADPH in cells. The cytoplasmic transhydrogenases could be useful to engineer synthetic biochemical pathways for the production of high-value chemicals and biofuels.Methodology/Principal FindingsA transhydrogenase activity was discovered for a FMN-bound diaphorase (DI) from Geobacillus stearothermophilus under anaerobic conditions. The DI-catalyzed hydride exchange were monitored and characterized between a NAD(P)H and a thio-modified NAD+ analogue. This new function of DI was demonstrated to transfer a hydride from NADPH to NAD+ that was consumed by NAD-specific lactate dehydrogenase and malic dehydrogenase.Conclusions/SignificanceWe discover a novel transhydrogenase activity of a FMN-DI by stabilizing the reduced state of FMNH2 under anaerobic conditions. FMN-DI was demonstrated to catalyze the hydride transfer between NADPH and NAD+. In the future, it may be possible to incorporate this FMN-DI into synthetic enzymatic pathways for balancing NADH generation and NADPH consumption for anaerobic production of biofuels and biochemicals.
Highlights
Cellular metabolism uses many cofactors for facilitating the electron transfer from one molecule to another in redox reactions
It may be possible to incorporate this flavin mononucleoide (FMN)-DI into synthetic enzymatic pathways for balancing NADH generation and Nicotinamide adenine dinucleotide phosphate (NADPH) consumption for anaerobic production of biofuels and biochemicals
Proton-translocating transhydrogenases are important in bioenergetics, where the hydride transfer from a NADH to a NADP+ is powered by an electrochemical proton gradient in mitochondria.[3,6]
Summary
Redox cofactors of NADH/NADPH participate in many cellular metabolic pathways for facilitating the electron transfer from one molecule to another in redox reactions. A transhydrogenase activity was discovered for a FMN-bound diaphorase (DI) from Geobacillus stearothermophilus under anaerobic conditions. The DI-catalyzed hydride exchange were monitored and characterized between a NAD(P)H and a thio-modified NAD+ analogue. This new function of DI was demonstrated to transfer a hydride from NADPH to NAD+ that was consumed by NAD-specific lactate dehydrogenase and malic dehydrogenase. It may be possible to incorporate this FMN-DI into synthetic enzymatic pathways for balancing NADH generation and NADPH consumption for anaerobic production of biofuels and biochemicals. A Hidden Transhydrogen Activity of a FMN-Bound Diaphorase under Anaerobic Conditions
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