Efficient NADPH regeneration based on co-expression of hexokinase and glucose-6-phosphate dehydrogenase

Abstract:

[Objective] This study aimed to construct a co-expression system of hexokinase and glucose-6-phosphate dehydrogenase in Escherichia coli to regenerate NADPH with glucose as substrate efficiently. [Methods] Hexokinase genes hkgs and hkpp were cloned and expressed in E. coli BL21(DE3), and then the co-expression system of hexokinase and glucose-6-phosphate dehydrogenase was established to achieve efficient in-situ regeneration of NADPH. Among the two co-expression systems, BL21(HKgs+GpdPP) was better, so the expression condition of BL21(HKgs+GpdPP) was optimized. [Results] The catalytic activity of NADPH regeneration reached 856 U/L. The coupling catalysis was performed with this co-enzyme regeneration system and alcohol dehydrogenase AdhR, the catalytic activity of asymmetric reduction of ethyl 4-chloro-3-oxobutanoate was enhanced up to 2.5 times. [Conclusion] Through co-expression system of hexokinase and glucose-6-phosphate dehydrogenase in Escherichia coli, new effective NADPH regeneration system was constructed, and success for a symmetric reduction by alcohol dehydrogenase.