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

Abstract

Background: 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. Results: 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 (μ max. = 0.31 h -1 ) and l-lysine productivity (q Lys, max. = 0.53 g g -1 h -1 ) as compared with strain RG (μ max. = 0.14 h -1 and q Lys, max. = 0.42 g g -1 h -1 ). Conclusions: 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.[Figure not available: see fulltext.]