Enantioselective Biosynthesis of L-Phenyllactic Acid From Phenylpyruvic Acid by L-Lactate Dehydrogenase Coupling With Glucose Dehydrogenase.

As a valuable versatile building block, L-phenyllactic acid (L-PLA) has numerous applications in the fields of agriculture, pharmaceuticals, and biodegradable plastics. However, both normally chemically synthesized and naturally occurring PLA are racemic, and the production titer of L-PLA is not satisfactory. To improve L-PLA production and reduce the high cost of NADH, an coenzyme regeneration system of NADH was achieved using the glucose dehydrogenase variant GDH and introduced into the L-PLA production process. Here an NADH-dependent L-lactate dehydrogenase-encoding variant gene (L-1) was expressed in GS115. The specific activity of L-LDH1 (Pp) was as high as 447.6 U/mg at the optimum temperature and pH of 40°C and 5.0, which was 38.26-fold higher than that of wild-type L-LDH1 (Pp). The catalytic efficiency ( / ) of L-LDH1 (Pp) was 94.3 mM s, which was 67.4- and 25.5-fold higher than that of L-LDH1(Pp) and L-LDH1 (Ec) expressed in , respectively Optimum reactions of L-PLA production by dual-enzyme catalysis were at 40°C and pH 5.0 with 10.0 U/ml L-LDH1 (Pp) and 4.0 U/ml GDH. Using 0.1 mM NAD, 400 mM (65.66 g/L) phenylpyruvic acid was completely hydrolyzed by fed-batch process within 6 h, affording L-PLA with 90.0% yield and over 99.9% . This work would be a promising technical strategy for the preparation of L-PLA at an industrial scale.