Understanding and Overcoming the Limitations of Bacillus badius and Caldalkalibacillus thermarum Amine Dehydrogenases for Biocatalytic Reductive Amination

• Published in: ACS catalysis 2017-05, Vol.7 (5), p.3204-3209
• Main Authors: Pushpanath, Ahir, Siirola, Elina, Bornadel, Amin, Woodlock, David, Schell, Ursula
• Format: Article
• Language: English
• ISSN: 2155-5435; 2155-5435
• DOI: 10.1021/acscatal.7b00516

The direct asymmetric reductive amination of ketones using ammonia as the sole amino donor is a growing field of research in both chemocatalysis and biocatalysis. Recent research has focused on the enzyme engineering of amino acid dehydrogenases (to obtain amine dehydrogenases), and this technology promises to be a potentially exploitable route for chiral amine synthesis. However, the use of these enzymes in industrial biocatalysis has not yet been demonstrated with substrate loadings above 80 mM, because of the enzymes’ generally low turnover numbers (k cat < 0.1 s–1) and variable stability under reaction conditions. In this work, a newly engineered amine dehydrogenase from a phenylalanine dehydrogenase (PheDH) from Caldalkalibacillus thermarum was recruited and compared against an existing amine dehydrogenase (AmDH) from Bacillus badius for both kinetic and thermostability parameters, with the former exhibiting an increased thermostability (melting temperature, T m) of 83.5 °C, compared to 56.5 °C for the latter. The recruited enzyme was further used in the reductive amination of up to 400 mM of phenoxy-2-propanone (c = 96%, ee (R) < 99%) in a biphasic reaction system utilizing a lyophilized whole-cell preparation. Finally, we performed computational docking simulations to rationalize the generally lower turnover numbers of AmDHs, compared to their PheDH counterparts.