X-ray structure of Escherichia coli pyridoxine 5′-phosphate oxidase complexed with pyridoxal 5′-phosphate at 2.0 Å resolution

Escherichia colipyridoxine 5′-phosphate oxidase catalyzes the terminal step in the biosynthesis of pyridoxal 5′-phosphate by the FMN oxidation of pyridoxine 5′-phosphate forming FMNH 2 and H 2O 2. Recent studies have shown that in addition to the active site, pyridoxine 5′-phosphate oxidase contains a non-catalytic site that binds pyridoxal 5′-phosphate tightly. The crystal structure of pyridoxine 5′-phosphate oxidase from E. coli with one or two molecules of pyridoxal 5′-phosphate bound to each monomer has been determined to 2.0 Å resolution. One of the pyridoxal 5′-phosphate molecules is clearly bound at the active site with the aldehyde at C4′ of pyridoxal 5′-phosphate near N5 of the bound FMN. A protein conformational change has occurred that partially closes the active site. The orientation of the bound pyridoxal 5′-phosphate suggests that the enzyme catalyzes a hydride ion transfer between C4′ of pyridoxal 5′-phosphate and N5 of FMN. When the crystals are soaked with excess pyridoxal 5′-phosphate an additional molecule of this cofactor is also bound about 11 Å from the active site. A possible tunnel exists between the two sites so that pyridoxal 5′-phosphate formed at the active site may transfer to the non-catalytic site without passing though the solvent.