Biotransformation of beta-keto nitriles to chiral (S)-beta-amino acids using nitrilase and omega-transaminase

Objective To enzymatically synthesize enantiomerically pure beta-amino acids from beta-keto nitriles using nitrilase and omega-transaminase. Results An enzyme cascade system was designed where in beta-keto nitriles are initially hydrolyzed to beta-keto acids using nitrilase from Bradyrhizobium japonicum and subsequently beta-keto acids were converted to beta-amino acids using omega-transaminases. Five different omega-transaminases were tested for this cascade reaction, To enhance the yields of beta-amino acids, the concentrations of nitrilase and amino donor were optimized. Using this enzymatic reaction, enantiomerically pure (S)-beta-amino acids (ee > 99%) were generated. As nitrilase is the bottleneck in this reaction, molecular docking analysis was carried out to depict the poor affinity of nitrilase towards beta-keto acids. Conclusions A novel enzymatic route to generate enantiomerically pure aromatic (S)-beta-amino acids from beta-keto nitriles is demonstrated for the first time

The Kinetic Resolution of Racemic Amines Using a Whole-Cell Biocatalyst Co-Expressing Amine Dehydrogenase and NADH Oxidase

Amine dehydrogenase (AmDH) possesses tremendous potential for the synthesis of chiral amines because AmDH catalyzes the asymmetric reductive amination of ketone with high enatioselectivity. Although a reductive application of AmDH is favored in practice, the oxidative route is interesting as well for the preparation of chiral amines. Here, the kinetic resolution of racemic amines using AmDH was first extensively studied, and the AmDH reaction was combined with an NADH oxidase (Nox) to regenerate NAD+ and to drive the reaction forward. When the kinetic resolution was carried out with 10 mM rac-2-aminoheptane and 5 mM rac-α-methylbenzylamine (α-MBA) using purified enzymes, the enantiomeric excess (ee) values were less than 26% due to the product inhibition of AmDH by ketone and the inhibition of Nox by the substrate amine. The use of a whole-cell biocatalyst co-expressing AmDH and Nox apparently reduces the substrate and product inhibition, and/or it increases the stability of the enzymes. Fifty millimoles (50 mM) rac-2-aminoheptane and 20 mM rac-α-MBA were successfully resolved into the (S)-form with >99% ee using whole cells. The present study demonstrates the potential of a whole-cell biocatalyst co-expressing AmDH and Nox for the kinetic resolution of racemic amines