NAD +-dependent formate dehydrogenase from the bacterium Staphylococcus aureus (SauFDH) plays an important role in the vital activity of these bacteria. The gene encoding SauFDH was successfully cloned and expressed in our laboratory. Since this enzyme has the highest k cat value among the described FDHs and also has a high temperature stability compared to other proteins, it can be considered as a promising catalyst for regeneration of NAD(P)H. The main disadvantage of this enzyme are high K M values. In this work, the principle of rational design was used to reduce K M. As a result, 9 catalytically significant positions 119, 194, 196, 217-219, 246, 303, and 323 were identified, and 16 new mutant forms of SauFDH were obtained and fully characterized. Substitutions in positions 119 and 194 lead to an increase of K M NAD+. In the Ile-Val-Ala-Gly line, position 119 tends to improve NAD + binding. K M NAD+ of SauFDH V119G decreased by 27 times compared to the wild-type enzyme. K M NAD+ Phe194Val decreased by 3.5 times. The catalytic constant for this mutant form practically did not change. The use of a complex buffer increased the catalytic efficiency 6 times for the mutant with the Phe194Val substitution compared to wt-SauFDH in a single-component buffer.