(280e) Extending the Utility of the Amine Dehydrogenase: Increased Affinity and Synthesis of (S)-Amine

Au, S., University of Kentucky
Bommarius, B., Georgia Institute of Technology
Abrahamson, M. J., Georgia Institute of Technology
Bommarius, A. S., Georgia Institute of Technology

The novel amine dehydrogenase (AmDH) has been recently developed1 and further characterized. Through protein engineering of the amino acid dehydrogenase scaffold, the amine dehydrogenase now catalyzes the reduction of prochiral ketones to chiral amines. Further protein engineering to the phenylalanine amine dehydrogenase (F-AmDH) to expand the binding pocket has led to an increased specific activity toward its model substrate, para fluoro phenyl acetone (pFPA). In addition, F-AmDH exhibited a low affinity for ammonia (KM: 550 mM), leading the enzyme to perform in unfavorable conditions. Protein engineering of residues in the binding pocket has led to an increased affinity towards ammonia.

Previous work involving the AmDH included the development of a biphasic organic solvent system to allow for conversion of hydrophobic substrates2. The F-AmDH naturally catalyzes formation of the (R)-amine. To further continue our reaction engineering work, oxidative amination in a biphasic organic solvent system has led to a successful conversion to the (S)-amine.  For proof of concept, we have produced (S)-methylbenzylamine ((S)-MBA) from the racemate, catalyzed by a chimeric (‘c’) AmDH.  We will discuss enantiomeric purity of the product, related degree of conversion, and reaction parameters such as residence time and enzyme loads.

[1] (a) Abrahamson, M. J.; Wong, J. W.; Bommarius, A. S., The Evolution of an Amine Dehydrogenase Biocatalyst for the Asymmetric Production of Chiral Amines. Adv Synth Catal 2013, 355 (9), 1780-1786; (b) Abrahamson, M. J.; Vazquez-Figueroa, E.; Woodall, N. B.; Moore, J. C.; Bommarius, A. S., Development of an Amine Dehydrogenase for Synthesis of Chiral Amines. Angew Chem Int Edit 2012, 51(16), 3969-3972; (c) Bommarius, B.; Schurmann, M.; Bommarius, A., A chimeric amine dehydrogenase offers surprising properties not found in either parent. 2014 (submitted).

[2] Samantha K. Au, B. R. B., Andreas S. Bommarius, Biphasic reaction system allows for conversion of hydrophobic substrates by amine dehydrogenases. 2014(revisions submitted)