Mechanism of Pyridoxine 5'-Phosphate Accumulation in Pyridoxal 5'-Phosphate-Binding Protein Deficiency

The pyridoxal 5'-phosphate (PLP)-binding protein (PLPBP) plays an important role in vitamin B₆ homeostasis. Loss of this protein in organisms such as Escherichia coli and humans disrupts the vitamin B₆ pool and induces intracellular accumulation of pyridoxine 5'-phosphate (PNP), which is normally undetectable in wild-type cells. This accumulated PNP could affect diverse metabolic systems through the inhibition of some PLP-dependent enzymes. In this study, we investigated the as-yet-unclear mechanism of intracellular accumulation of PNP due to the loss of PLPBP protein encoded by yggS in E. coli. Genetic studies using several PLPBP-deficient strains of E. coli lacking a known enzyme(s) in the de novo or salvage pathways of vitamin B₆, including pyridoxine (amine) 5'-phosphate oxidase (PNPO), PNP synthase, pyridoxal kinase, and pyridoxal reductase, demonstrated that neither the flux from the de novo pathway nor the salvage pathway solely contributed to the PNP accumulation caused by the PLPBP mutation. Studies of the strains lacking both PLPBP and PNPO suggested that PNP shares the same pool with PMP, and showed that PNP levels are impacted by PMP levels and vice versa. Here, we show that disruption of PLPBP perturbs PMP homeostasis, which may result in PNP accumulation in the PLPBP-deficient strains. IMPORTANCE A PLP-binding protein (PLPBP) from the conserved COG0325 family has recently been recognized as a key player in vitamin B₆ homeostasis in various organisms. Loss of PLPBP disrupts vitamin B₆ homeostasis and perturbs diverse metabolisms, including amino acid and α-keto acid metabolism. Accumulation of PNP is a characteristic phenotype of PLPBP deficiency and is suggested to be a potential cause of the pleiotropic effects, but the mechanism of this accumulation has been poorly understood. In this study, we show that fluxes for PNP synthesis/metabolism are not responsible for the accumulation of PNP. Our results indicate that PLPBP is involved in the homeostasis of pyridoxamine 5'-phosphate, and that its disruption may lead to the accumulation of PNP in PLPBP deficiency.