Engineering Coenzyme Specificity of Formate Dehydrogenases: The Role of Amino Acid Residues at Positions 379 and 380

The mechanism of selective specificity of oxidoreductases to NAD+ or NADP+ and the ability to change the coenzyme specificity of these enzymes are some of the most important fundamental and applied problems. The first work on the switch in the coenzyme specificity from NADP+ to NAD+ was performed in 1990 for glutathione reductase. In 1993, formate dehydrogenase (FDH, EC 1.2.1.2) from the methylotrophic bacterium Pseudomonas sp. 101 (PseFDH) became the first oxidoreductase whose coenzyme specificity was changed in the opposite direction – from NAD+ to NADP+. Mutant NADP+-specific FDHs are extensively used in fine organic synthesis (including production of chiral compounds). The switch in the coenzyme specificity from NAD+ to NADP+ in FDHs is achieved by substituting amino acids at positions 198, 221, 222, 260, 379, and 380 (numbering according to PseFDH); however available data do not allow the interpretation of the exact role of each individual substitution. Since 2010, five natural NADP+-dependent FDHs have been found. In 2015-2024, three 3D structures for two natural and four 3D structures for two mutant NADP+-specific FDHs have appeared in the Protein Data Bank (PDB). In this review, we briefly discussed the general principles of coenzyme specificity based on the experimental and modeled FDH structures and performed a detailed analysis of the type and arrangement of residues at positions corresponding to His379 and Ser380 in PseFDH, whose role in NADP+ binding is still debated.