Enzymatic Preparation of an (S)-Amino Acid from a Racemic Amino Acid

The ( S )-amino acid, ( S )-2-amino-3-(6- o -tolylpyridin-3-yl)propanoic acid ( 3 ), is a key intermediate needed for synthesis of an antidiabetic drug candidate. Three enzymatic routes to 3 were explored. ( S )-Amino acid 3 could be prepared in 73% isolated yield with 99.9% ee from racemic amino acid 1 using ( R )-amino acid oxidase from Trigonopsis variabilis expressed in Escherichia coli in combination with an ( S )-aminotransferase using ( S )-aspartate as amino donor. The ( S )-aminotransferase was purified from a soil organism identified as Burkholderia sp. and cloned and expressed in E. coli . ( S )-Amino acid 3 with 100% ee was also prepared in 68% solution yield and 54% isolated yield from 1 using recombinant ( R )-amino acid oxidase from T. variabilis and an ( S )-amino acid dehydrogenase from Sporosarcina ureae . The cofactor NADH required for the reductive amination reaction was regenerated using formate and formate dehydrogenase. The chemoenzymatic dynamic resolution of 1 by ( R )-selective oxidation with Celite-immobilized ( R )-amino acid oxidase in combination with chemical imine reduction using borane−ammonia complex gave an 81% solution yield and 68% isolated yield of 3 with 100% ee.