Engineering of NADPH regenerators in Escherichia coli for enhanced biotransformation

Abstract

Efficient regeneration of NADPH is one of the limiting factors that constrain the productivity of biotransformation processes. In order to increase the availability of NADPH for enhanced biotransformation by engineered Escherichia coli, modulation of the pentose phosphate pathway and amplification of the transhydrogenases system have been conventionally attempted as primary solutions. Recently, other approaches for stimulating NADPH regeneration during glycolysis, such as replacement of native glyceradehdye-3-phosphate dehydrogenase (GAPDH) with NADP-dependent GAPDH from Clostridium acetobutylicum and introduction of NADH kinase catalyzing direct phosphorylation of NADH to NADPH from Saccharomyces cerevisiae, were attempted and resulted in remarkable impacts on NADPH-dependent bioprocesses. This review summarizes several metabolic engineering approaches used for improving the NADPH regenerating capacity in engineered E. coli for whole-cell-based bioprocesses and discusses the key features and progress of those attempts.

NADP + -Preferring d -Lactate Dehydrogenase from Sporolactobacillus inulinus

“…The energy-independent, soluble transhydrogenase UdhA of E. coli has been reported to convert NADPH and NAD ϩ to NADP ϩ and NADH, respectively, under high concentrations of NADPH (31). It is of interest to determine whether DLDH744 had a similar transhydrogenase activity and therefore used regenerated NADH as a coenzyme.…”

“…Therefore, no hydrogen was transferred from NADPH to NAD ϩ to produce NADH. Additionally, NADH kinases were reported to directly phosphorylate NADH to produce NADPH, which subsequently enhanced the productivity of the NADPHdependent biotransformation processes (31). To exclude the possibility that NADH was being produced from NADPH by reverse NADH kinase activity, the quantity of released phosphate was measured using Milli-Q water in the enzymatic reaction mixture.…”

Thermococcus kodakarensis : the key to affordable biohydrogen production

“…Not only is NADPH vital for the generation of reactive oxygen species (ROS) [111][112][113][114] and the anti-oxidative defense mechanisms of most organisms [115][116][117], most importantly, it is also the driving force of most biosynthetic enzymatic reactions, including those responsible for the biosynthesis of all major cell components, such as DNA and lipids [115][116][117][118][119][120][121]. Owing to this essential role in biosynthesis, NADPH availability has been of major interest to industry [122,123]. Many natural products of industrial importance are complex secondary metabolites, the production of which often involves NADPH-dependent enzymes.…”

“…They also contain numerous pathways, involving stable and highly specific enzymes, thus obviating the need for expensive enzyme purification. In addition, our knowledge of natural metabolic pathways is rapidly advancing, allowing for rational design towards product formation [122,123,133,134]. Therefore, it is not surprising that microbial conversion is the preferred method for the synthesis of a range of products.…”

“…This suggests that, under the applied conditions, IDH is not a better option than G6PDH for NADPH regeneration in biotransformation reactions [122]. The ability of IDH overexpression to enhance glutamate formation in Corynebacterium glutamicum has also been tested [247].…”