Structural and Functional Insights into the Escherichia Coli Glycerol Dehydrogenase

Abstract

During glycerol metabolism, the initial step of glycerol oxidation is catalyzed by glycerol dehydrogenase (GDH), which converts glycerol to dihydroxyacetone in a NAD+-dependent manner via an ordered Bi-Bi kinetic mechanism. Structural studies conducted with GDH from various species have mainly elucidated structural details of the active site and ligand binding. However, the structure of the full GDH complex with both cofactor and substrate bound is not determined, and thus, the structural basis of the kinetic mechanism of GDH remains unclear. Here, I report the crystal structures of Escherichia coli GDH with a substrate analogue bound in the absence or presence of NAD+. Structural analyses including molecular dynamics simulations revealed that GDH possesses a flexible β-hairpin, and that during the ordered progression of the kinetic mechanism, the flexibility of the β-hairpin is reduced after NAD+ binding. It was also observed that this alterable flexibility of the β-hairpin contributes to the cofactor binding and possibly to the catalytic efficiency of GDH. In addition, I report another crystal structure of E. coli GDH complexed with NAD+ and glycerol, and analyze GDH’s octameric oligomerization, suggesting its potential role for the enzyme activity. These studies expand our understanding of the GDH structure and its relation to the enzymatic activity.