Characterization of bacterial l-galactose dehydrogenase with l-glucose dehydrogenase activity from Luteolibacter sp....

The crystal structure of l-galactose dehydrogenase (LGDH) with l-glucose dehydrogenase activity from Luteolibacter sp. strain LG18 (Lu-LGDH) was determined in complex with l-galactose or l-glucose and NADP⁺. This structural analysis identified key residues involved in substrate binding, and alanine-substituted mutants demonstrated the roles of these residues, including Tyr56, acting as a potential general base within the catalytic tetrad. Unlike plant enzymes that show a preference for NAD⁺, Lu-LGDH exhibits a marked preference for NADP⁺ as a cofactor. This preference was attributed to the interaction of the phosphate group with Arg28, Thr269, and Asn274. The binding mode of l-glucose was similar to that of l-galactose. The C4 hydroxyl group (the structural difference between these pyranoses) was not used for substrate binding, which explains the dual activity of the enzyme. Furthermore, among the substrate-binding residues that were mutated, Arg308, which is not conserved among LGDHs, was crucial for the enzymatic activity.