The production of single enantiomers of chiral intermediates has become increasingly important in the pharmaceuticals.  Chiral intermediates were prepared for the synthesis of Saxagliptin, a dipeptidyl peptidase IV inhibitor under development for treatment of type 2 diabetes mellitus. The amino acid 2-(3-hydroxy-1-adamantyl)-(2S)-aminoethanoic acid, [(S)-3-hydroxyadamantylglycine], was prepared from keto acid 2-(3-hydroxy-1-adamantyl)-2-oxoethanoic acid by reductive amination using a phenylalanine dehydrogenase from Thermoactinomyces intermedius expressed in Pichia pastoris or Escherichia coli. NAD produced during the reaction was recycled to NADH using formate dehydrogenase. Pichia pastoris produces an endogenous formate dehydrogenase when grown on methanol, and the corresponding gene was cloned and expressed in E. coli. Production of multi-kg batches was carried out with extracts of Pichia pastoris expressing the modified phenylalanine dehydrogenase from Thermoactinomyces intermedius and endogenous formate dehydrogenase, and further scaled up using a preparation of the two enzymes expressed in E. coli.
An efficient biocatalytic method has been developed for the conversion of (5S)-4,5-dihydro-1H-pyrrole-1,5-dicarboxylic acid, 1-(1,1-dimethylethyl)-5-ethyl ester into the corresponding amide (5S)-5-aminocarbonyl-4,5-dihydro-1H-pyrrole-1-carboxylic acid, 1-(1,1-dimethylethyl)ester. Candida antartica lipase B mediates ammonolysis of the ester with ammonium carbamate as ammonia donor to yield up to 98% of the amide, offering an efficient and practical alternative to chemical routes.