Unlocking Catalytic Diversity of a Formate Dehydrogenase: Formamide Activity for NADPH Regeneration and Amine Supply for Asymmetric Reductive Amination
Artur Maier, Tanja Knaus, Francesco G. Mutti, Dirk Tischler
Abstract
High Resolution Image Download MS PowerPoint Slide The formate dehydrogenase (FDH) from Candida boidinii is a well-studied and applied enzyme for NADH regeneration in various reactions. As many oxidoreductases require NADPH, FDH mutants were created with shifted cofactor specificity toward NADP + . However, less effort was made to elucidate the substrate specificity for the hydride donors. Here, we report the FDH-catalyzed cleavage of formamide (F) and derivatives thereof into CO 2 and amines, while regenerating the cofactors NADH and NADPH. Wild-type FDH and the NADP + -accepting variant FDH C23S/D195Q/Y196R/Q197N (FDH M5) showed both activity with 10% (v/v) F, N -methylformamide (MF), and N, N -dimethylformamide of 80, 67, and 4.5 mU/mg, and 4.9, 4.7, and 0.5 mU/mg, respectively. In silico docking and molecular dynamics simulation gave insights into substrate binding, indicating an altered binding conformation. NADP + -accepting variants were utilized in a cascade set up for the reductive amination of cyclohexanone by means of reductive aminase from Aspergillus oryzae with MF as hydride and amine donor, thereby reaching conversion rates of 72% in a whole cell approach. This work broadens the applicability of FDHs in biocatalysis.