Development of an NAD(H)‐Driven Biocatalytic System for Asymmetric Synthesis of Chiral Amino Acids

Abstract Chiral amino acids are extensively applied in the pharmaceutical, food, cosmetic, and agricultural industries. As a representative example, l ‐phosphinothricin ( l ‐PPT, a chiral non‐natural amino acid) is a broad‐spectrum herbicide. An NAD(H)‐driven biocatalytic system for the asymmetric synthesis of chiral amino acids (focused on l ‐PPT) with high efficiency and low cost is highly desired. The key for the development of such biocatalytic system is to obtain an NADH‐dependent biocatalyst with high catalytic performance toward l ‐PPT pro‐ketone PPO. Herein, an engineered glutamate dehydrogenase from Lysinibacillus composti ( Lc GluDH) with desired activity was obtained by gene mining and protein engineering. In silico analyses suggested that the volume of substrate‐binding pocket was substantially enlarged from 330.5 Å 3 to 409.6 Å 3 . The stability of Lc GluDH was increased ( T m value increased from 47.3 °C to 65.3 °C) by introducing positively charged amino acids or aromatic amino acids at position 375. The desired biocatalytic system was constructed by coupling the engineered Lc GluDH and an NAD + ‐dependent FDH. Through this biocatalytic system, the batch production of l ‐PPT exhibited high space‐time yield (207.3 g ⋅ L −1 ⋅ day −1 ) with strict stereoselectivity ( ee of l ‐PPT>99%). Furthermore, eight other chiral amino acids were synthesised by the developed NAD(H)‐driven biocatalytic system with high ee values. magnified image