Production of L-Lactic Acid in Saccharomyces cerevisiae Through Metabolic Engineering and Rational Cofactor Engineering

Microbial engineering based on synthetic biology can facilitate large-scale production of target products. In this study, the introduction of lactate dehydrogenase (LDH) enabled Saccharomyces cerevisiae to acquire the capacity for L-lactic acid (LA) production and the NADH/NAD + ratio from 0.228 to 0.156, while the subsequent modification of carbon metabolism pathway led to a rapid increase of NADH/NAD + even up to 0.337. By testing the effectiveness of four different redox systems, we demonstrated that dynamic regulation of additional redox genes to consume excessive NADH is more beneficial for LA accumulation, alleviating the negative effects of metabolic modification on hosts, and altering the distribution of metabolic flow. We first reported expression of GLT1 which coding glutamate synthase has the strongest ability to increase LA production and reduce NADH/NAD + . Combining metabolic engineering and cofactor engineering, the LA yield reached from 0.04 g/g to 0.37 g/g in YNB medium. Subsequently, strain PK27 produced 37.94 g/L LA with production yield of 0.66 g/g in YPD medium. Finally, the results could provide a reference that the potential under poor nutrient culture conditions and the direction and intensity of regulation of intracellular NADH/NAD + for LA accumulation.