A combination fermentation strategy for simultaneously increasing cellular NADP(H) level, biomass, and enzymatic activity of glufosinate dehydrogenase in Escherichia coli

Oxidoreductase is one of the most important biocatalysts for the synthesis of various chiral compounds. However, their whole-cell activity is frequently affected by an insufficient supply of expensive nicotinamide cofactors. This study aimed to overcome such shortcomings by developing a combination fermentation strategy for simultaneously increasing intracellular NADP(H) level, biomass, and glufosinate dehydrogenase activity in E. coli . The results showed that the feeding mode of NAD(H) synthesis precursor and lactose inducer had essential effects on the accumulation level of intracellular NADPH. Adding 40 mg L −1 of L-aspartic acid to the medium increased the intracellular NADP(H) concentration by 36.3%. Under the pH-stat feeding mode and adding 0.4 g L −1 h −1 lactose, the NADP(H) concentration, biomass, and GluDH activity in the 5-L fermenter reached 445.7 μmol L −1 , 21.7 gDCW L −1 , and 8569.3 U L −1 , respectively. As far as we know, this is the highest reported activity of GluDH in the fermentation broth. Finally, the 5000-L fermenter was successfully scaled up to use this fermentation approach. The combination fermentation strategy might serve as a useful approach for the high-activity fermentation of other NADPH-dependent oxidoreductases.