Amine-Tolerant E. coli Strains Generated <i>via</i> Adaptive\nEvolution for Sustainable\nSynthesis of Chiral Amines

The\nbiocatalytic synthesis of chiral amines from carbonyl\ncompounds\nand ammonia is a major advance in sustainable synthetic chemistry.\nUsing whole cells for bioamination reactions is advantageous given\ntheir low preparation cost and direct applicability; however, amine\ntoxicity limits the reaction when living cells are used. Herein, we\nadapted Escherichia coli BL21(DE3)\ncells to grow in the presence of 100 mM hexan-2-amine <i>via</i> directed evolution in continuous culture, obtaining six times more\ntolerant strains than the wild-type. The adapted strains also displayed\nsuperior tolerance for structurally different amines. Coexpression\nof genes encoding for amine dehydrogenase (AmDH) and formate dehydrogenase\nactivities in the adapted strains enabled the stereoselective bioamination\n(ee > 99%) of different prochiral ketones with up to 80% conversion\nat high substrate loading (up to 200 mM) without exogenous cofactor\naddition. The adapted cells displayed longer survival and higher population\ndensity during the reactions. The present biotechnological E. coli system contributes to the development of\nmore robust biocatalysis for amine production.