QueF nitrile reductases: Catalytic mechanism and advances in biocatalyst development

Nitrile-to-amine reduction is a key transformation in organic and industrial synthesis, providing access to pharmaceutical, agrochemical, and materials manufacturing relevant primary amines. QueF nitrile reductases represent a unique class of biocatalysts capable of catalyzing the NAD(P)H-dependent four-electron reduction of nitriles under mild conditions. Despite their biocatalytic potential, the practical application of QueF enzymes is severely constrained by an exceptionally narrow substrate scope. This review presents a comprehensive analysis of QueF research, from its discovery, structural and mechanistic elucidation to the most recent protein-engineering efforts. Experimental evidence indicates that the long evolutionary optimization of the active site, characterized by a highly specialized substrate-binding pocket, is the primary determinant of QueF's substrate specificity. A critical analysis of the relationships between key active-site residues and substrate functional groups, integrating structural, mutagenesis, and computational insights, rationalizes selectivity trends. Ultimately, this work outlines the fundamental challenges associated with expanding QueF substrate tolerance and identifies plausible strategies for future engineering. Continued progress will rely on a deeper understanding of catalytic mechanism, active-site dynamics, and coordinated multi-residue redesign, positioning QueF as a valuable model system for the development of next-generation nitrile-reducing biocatalysts.