Biocatalytic asymmetric synthesis of L-phosphinothricin using a one-pot three enzyme system and a continuous substrate fed-batch strategy

Transamination catalyzed by an aminotransferase is becoming a key tool for the production of chiral amine pharmaceuticals and agrochemicals owing to its excellent enantioselectivity and green credentials. To overcome the unfavorable thermodynamic equilibrium and the inhibition by the byproduct alpha-ketoglutaric acid in the transamination of 2-oxo-4-[(hydroxy)(methyl)phosphinoyl)butyric acid (PPO) to form the promising herbicide L-phosphinothricin (L-PPT), a tri-enzymatic cascade reaction system was developed by combining a robust glutamate dehydrogenase to recycle the byproduct to the amino donor L-glutamate in situ, together with a cofactor recycling process catalyzed by an alcohol dehydrogenase. Moreover, a continuous substrate fed-batch strategy was employed to alleviate the decomposition of PPO and applied to scale-up the cascade reaction to 90 L, yielding 111.4 g/L (615.4 mM) L-PPT in 99.7% yield and > 99.9% ee with an productivity of 15.9 g/L.h. This combination of improved biocatalyst system and process engineering should prove to be economically competitive for industrial applications.