X-ray structure of Escherichia coli pyridoxine 5 '-phosphate oxidase complexed with FMN at 1.8 angstrom resolution

Background: Escherichia coli pyridoxine 5'-phosphate oxidase (PNPOx) cataly zes the terminal step in the biosynthesis of pyridoxal 5'-phosphate (PLP), a cofactor used by many enzymes involved in amino acid metabolism. The enzy me oxidizes either the 4'-hydroxyl group of pyridoxine 5'-phosphate (PNP) o r the 4'-primary amine of pyridoxamine 5'-phosphate (PMP) to an aldehyde. P NPOx is a homodimeric enzyme with one flavin mononucleotide (FMN) molecule non-covalently bound to each subunit. A high degree of sequence homology am ong the 15 known members of the PNPOx family suggests that all members of t his group have similar three-dimensional folds. Results: The crystal structure of PNPOx from E. coli has been determined to 1.8 Angstrom resolution. The monomeric subunit folds into an eight-strande d beta sheet surrounded by five alpha-helical structures. Two monomers rela ted by a twofold axis interact extensively along one-half of each monomer t o form the dimer. There are two clefts at the dimer interface that are symm etry-related and extend from the top to the bottom of the dimer, An FMN cof actor that makes interactions with both subunits is located in each of thes e two clefts. Conclusions: The structure is quite similar to the recently deposited 2.7 A ngstrom structure of Saccharomyces cerevisiae PNPOx and also, remarkably, s hares a common structural fold with the FMN-binding protein from Desulfovib rio vulgaris and a domain of chymotrypsin. This high-resolution E. coli PNP Ox structure permits predictions to be made about residues involved in subs trate binding and catalysis. These predictions provide testable hypotheses, which can be answered by making site-directed mutants.