Engineered polypeptides that exhibit increased catalytic efficiency for unnatural cofactors and uses thereof

An engineered polypeptide having dehydrogenase activity that exhibits 100-fold or more catalytic efficiency for a nicotinamide-based unnatural cofactor relative to a wild-type or parent polypeptide that encodes a dehydrogenase,

• wherein the engineered polypeptide comprises 1, 2, 3, 4, 5, 6, 7, or 8 introduced amino acid substitutions in comparison to the sequence of the wild-type or parent polypeptide,
• wherein the introduced amino acid substitutions increase electrostatic complementarity between the engineered polypeptide and the unnatural cofactor,
• wherein the nicotinamide-based unnatural cofactor is a cofactor that is not normally utilized by the dehydrogenase encoded by the wild-type or parent polypeptide to catalyze a reaction, and
• wherein the cofactor normally utilized by the wild-type or parent polypeptide is nicotinamide adenine dinucleotide (NAD+) or nicotinamide adenine dinucleotide phosphate (NADP+).