Expanding the Substrate Scope of Native Amine Dehydrogenases through <i>In Silico</i> Structural Exploration and Targeted Protein Engineering

Native Amine Dehydrogenases (nat-AmDHs) are NAD(P)H-enzymes performing reductive amination, mainly active towards small aliphatic aldehydes and cyclic ketones, due to site volumes limited by the presence of several bulky amino acids. Herein, inspired diversity residues at these positions among family, we report implementation mutations F140A W145A in CfusAmDH their transposition nine other members. Moderate high conversions were obtained with substrates not accepted native enzymes, notably n-alkylaldehydes (44.6 %–99.5 % for hexanal nonanal) n-alkylketones (16.0 %–53.7 hexan-2-one nonan-2-one) retention excellent (S)-enantioselectivity (>99 ee). Complementary reported (R)-selective AmDHs, promising mutant CfusAmDH−W145A was further characterized its synthetic potential. Crystal structure resolution molecular dynamics gave insights into cofactor substrate specificity whole structural dynamics, thus providing keys mutagenesis work on this enzyme family.