Efficient synthesis of (R)-4-methoxyamphetamine and its analogues under low ammonium concentration using engineered amine dehydrogenase

Amphetamine and its analogues are widely used in various drugs because of its powerful and diverse effects on the nervous system of the brain, e.g., (R)-4-methoxyamphetamine is the key intermediate in formoterol for the treatment of asthma. Biocatalytic reductive amination of ketone by amine dehydrogenase (AmDH) is a potential method for the synthesis of chiral primary amines, but usually requires a high ammonium concentration to drive the amination reaction. In this study, iterative evolution was performed to generate a LsAmDH mutant W5-5 (T123G/S157T/S275G), which was able to catalyze the reductive amination of 200 mM 4-methoxypropiophe-none at 500 mM ammonium concentration to afford (R)-4-methoxyamphetamine with >99% ee and 90% iso-lated yield. This variant also catalyzed the reductive amination of other phenylacetone analogues with high yields and excellent enantioselectivity at low ammonium concentration. This would greatly reduce the genera-tion of ammonium-containing wastes and make the reaction more economic and greener, laying a foundation for the industrial application of AmDHs.