The chemical processes taking place in humans intersects the myriad of metabolic pathways occurring in commensal microorganisms that colonize the body to generate a complex biochemical network that regulates multiple aspects of human life. The role of tryptophan (Trp) metabolism at the intersection between the host and microbes is increasingly being recognized, and multiple pathways of Trp utilization in either direction have been identified with the production of a wide range of bioactive products. It comes that a dysregulation of Trp metabolism in either the host or the microbes may unbalance the production of metabolites with potential pathological consequences. The ability to redirect the Trp flux to restore a homeostatic production of Trp metabolites may represent a valid therapeutic strategy for a variety of pathological conditions, but identifying metabolic checkpoints that could be exploited to manipulate the Trp metabolic network is still an unmet need. In this review, we put forward the hypothesis that pyridoxal 5′-phosphate (PLP)-dependent enzymes, which regulate multiple pathways of Trp metabolism in both the host and in microbes, might represent critical nodes and that modulating the levels of vitamin B6, from which PLP is derived, might represent a metabolic checkpoint to re-orienteer Trp flux for therapeutic purposes.

Peroxisomal matrix proteins are transported into peroxisomes in a fully-folded state, but whether multimeric proteins are imported as monomers or oligomers is still disputed. Here, we used alanine:glyoxylate aminotransferase (AGT), a homodimeric pyridoxal 5′-phosphate (PLP)-dependent enzyme, whose deficit causes primary hyperoxaluria type I (PH1), as a model protein and compared the intracellular behavior and peroxisomal import of native dimeric and artificial monomeric forms. Monomerization strongly reduces AGT intracellular stability and increases its aggregation/degradation propensity. In addition, monomers are partly retained in the cytosol. To assess possible differences in import kinetics, we engineered AGT to allow binding of a membrane-permeable dye and followed its intracellular trafficking without interfering with its biochemical properties. By fluorescence recovery after photobleaching, we measured the import rate in live cells. Dimeric and monomeric AGT displayed a similar import rate, suggesting that the oligomeric state per se does not influence import kinetics. However, when dimerization is compromised, monomers are prone to misfolding events that can prevent peroxisomal import, a finding crucial to predicting the consequences of PH1-causing mutations that destabilize the dimer. Treatment with pyridoxine of cells expressing monomeric AGT promotes dimerization and folding, thus, demonstrating the chaperone role of PLP. Our data support a model in which dimerization represents a potential key checkpoint in the cytosol at the crossroad between misfolding and correct targeting, a possible general mechanism for other oligomeric peroxisomal proteins.