Equilibrium of the intracellular redox state for improving cell growth and l-lysine yield of Corynebacterium glutamicum by optimal cofactor swapping

NAD(H/+) and NADP(H/+) are the most important redox cofactors in bacteria. However, the intracellular redox balance is in advantage of the cell growth and production of NAD(P)H-dependent products. In this paper, we rationally engineered glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and isocitrate dehydrogenase (IDH) to switch the nucleotide-cofactor specificity resulting in an increase in final titer [from 85.6 to 121.4 g L−1] and carbon yield [from 0.33 to 0.46 g (g glucose)−1] of l-lysine in strain RGI in fed-batch fermentation. To do this, we firstly analyzed the production performance of original strain JL-6, indicating that the imbalance of intracellular redox was the limiting factor for l-lysine production. Subsequently, we modified the native GAPDH and indicated that recombinant strain RG with nonnative NADP-GAPDH dramatically changed the intracellular levels of NADH and NADPH. However, l-lysine production did not significantly increase because cell growth was harmed at low NADH level. Lastly, the nonnative NAD-IDH was introduced in strain RG to increase the NADH availability and to equilibrate the intracellular redox. The resulted strain RGI showed the stable ratio of NADPH/NADH at about 1.00, which in turn improved cell growth (μmax. = 0.31 h−1) and l-lysine productivity (qLys, max. = 0.53 g g−1 h−1) as compared with strain RG (μmax. = 0.14 h−1 and qLys, max. = 0.42 g g−1 h−1). This is the first report of balancing the intracellular redox state by switching the nucleotide-cofactor specificity of GAPDH and IDH, thereby improving cell growth and l-lysine production.

Abstract licence: CC BY-NC-ND

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   NADPH-generating systems in bacteria and archaea · 2015

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