Pdx3p is the pyridoxine/pyridoxamine 5'-phosphate oxidase (PNPOx; EC 1.4.3.5) of Saccharomyces cerevisiae, an FMN-containing homodimeric flavoenzyme that oxidizes pyridoxine 5'-phosphate and pyridoxamine 5'-phosphate to the cofactor pyridoxal 5'-phosphate (PLP), with production of hydrogen peroxide. It is the oxidase step of the vitamin B6 salvage pathway, acting after the kinase Bud16p; de novo PLP is made separately by the Snz1/Sno1 synthase. pdx3 mutants lack PNP/PMP oxidase activity, cannot use pyridoxine as a B6 source, and show pleiotropic defects in amino acid, heme, sterol and fatty acid metabolism that follow from PLP shortage. Recombinant Pdx3p binds FMN and is catalytically active, whereas the distantly related yeast proteins Ylr456w and Ypr172w, which share only the N-terminal PNPOx domain, are inactive. Pdx3p has been detected in the mitochondrial intermembrane space.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004733 pyridoxamine phosphate oxidase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PANTHER PTN008507293, seeded by characterized PNPOx enzymes (E. coli PdxH, human PNPO, Arabidopsis, fly, mouse, rat, zebrafish) and by Pdx3 itself. Pdx3 has the full two-domain PNPOx architecture and confirmed activity, so the inheritance is sound. Reason: GO:0004733 is defined as PMP + H2O + O2 = PLP + NH4+ + H2O2 and also covers oxidation of PNP to PLP + H2O2, so it captures both Pdx3 reactions. PTN008507293 is the Bacteria-rooted PNPOx node of PTHR10851; Pdx3 is in the SF0 subfamily that inherited it, retains both PNPOx domains, and its own direct assay is among the seeds (expected, not circular). No PAINT loss on this lineage. Supporting Evidence: file:interpro/panther/PTHR10851/PTHR10851-paint.tsv PTHR10851 PTN008507293 GO:0004733 F IBD false PMID:26327315 On the other hand, Pdx3 recombinant protein exhibits all expected properties of a bona fide P(N/M)P oxidase. |
| GO:0004733 pyridoxamine phosphate oxidase activity | IDA PMID:26327315 Experimental Evidence for a Revision in the Annotation of Pu... | ACCEPT | Summary: IDA from Domitrovic et al. 2015, where recombinant Pdx3 oxidised PNP to PLP in vitro, in contrast to the inactive Ylr456w/Ypr172w. Reason: GO:0004733 is defined as PMP + H2O + O2 = PLP + NH4+ + H2O2 and also covers oxidation of PNP to PLP + H2O2, so it captures both Pdx3 reactions. Recombinant Pdx3 bound FMN and formed PLP, unlike the single-domain Ylr456w/Ypr172w. Supporting Evidence: PMID:26327315 On the other hand, Pdx3 recombinant protein exhibits all expected properties of a bona fide P(N/M)P oxidase. |
| GO:0004733 pyridoxamine phosphate oxidase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined IEA from InterPro IPR000659 plus RHEA:15149/15817 and EC 1.4.3.5 on the UniProt entry; matches the experimentally shown activity. Reason: GO:0004733 is defined as PMP + H2O + O2 = PLP + NH4+ + H2O2 and also covers oxidation of PNP to PLP + H2O2, so it captures both Pdx3 reactions. The UniProt reactions match the experimentally shown activity. Supporting Evidence: file:yeast/PDX3/PDX3-uniprot.txt Reaction=pyridoxine 5'-phosphate + O2 = pyridoxal 5'-phosphate + H2O2; |
| GO:0004733 pyridoxamine phosphate oxidase activity | IMP PMID:7896706 Sterol uptake induced by an impairment of pyridoxal phosphat... | ACCEPT | Summary: IMP from Loubbardi et al. 1995 (abstract cached); aux30/pdx3 mutants lack P(N/M)P oxidase activity in extracts and the cloned PDX3 gene complements. Reason: GO:0004733 is defined as PMP + H2O + O2 = PLP + NH4+ + H2O2 and also covers oxidation of PNP to PLP + H2O2, so it captures both Pdx3 reactions. Mutants lacking PNP/PMP oxidase activity in extracts, complemented by cloned PDX3, support the activity genetically; the curator read the full text. Supporting Evidence: PMID:7896706 These mutants are characterized by a lack in pyridoxine (pyridoxamine) phosphate oxidase [P(N/M)P oxidase] (EC 1.4.3.5) activity. |
| GO:0004733 pyridoxamine phosphate oxidase activity | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways PLPSAL-PWY assigns Pdx3 to the EC 1.4.3.5 PNP and PMP oxidation steps. Correct. Reason: GO:0004733 is defined as PMP + H2O + O2 = PLP + NH4+ + H2O2 and also covers oxidation of PNP to PLP + H2O2, so it captures both Pdx3 reactions. The YeastPathways salvage model assigns Pdx3 to both EC 1.4.3.5 steps. Supporting Evidence: PMID:26327315 On the other hand, Pdx3 recombinant protein exhibits all expected properties of a bona fide P(N/M)P oxidase. |
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN000091367 seeded by Pdx3 (SGD:S000000239) and Arabidopsis AT5G49970. Consistent with detection of Pdx3 in the mitochondrial intermembrane space; a more precise IMS annotation exists. Reason: Root-organelle IBA checked against the gene: the IBD node PTN000091367 is seeded by Pdx3 itself and Arabidopsis AT5G49970, and Pdx3 was identified in the mitochondrial intermembrane-space proteome, so the organelle claim holds for this protein. The IMS row is the sub-compartment; the organelle row is kept beside it, as the organelle policy requires. Supporting Evidence: file:interpro/panther/PTHR10851/PTHR10851-paint.tsv PTHR10851 PTN000091367 GO:0005739 C IBD false AGI_LocusCode:AT5G49970|SGD:S000000239 PMID:22984289 YBR035CPDX3Pyridoxine phosphate oxidase |
| GO:0005758 mitochondrial intermembrane space | IDA PMID:22984289 Intermembrane space proteome of yeast mitochondria. | ACCEPT | Summary: IDA from the IMS proteome (Vogtle et al. 2012), which identified Pdx3 among Bax-released intermembrane-space proteins; its localization was previously listed as unknown. Whether a cytosolic pool also exists is unclear. Reason: Pdx3 was among proteins released from mitochondria by Bax-mediated outer-membrane permeabilization and quantified by SILAC; its earlier localization was listed as unknown. Direct evidence; the assay resolves the IMS from the matrix. Supporting Evidence: PMID:22984289 YBR035CPDX3Pyridoxine phosphate oxidase |
| GO:0005758 mitochondrial intermembrane space | IEA GO_REF:0000044 | ACCEPT | Summary: SubCell mapping of the UniProt location, which derives from the same IMS proteome study. Reason: Electronic mapping of the UniProt subcellular location, itself derived from PMID:22984289. Supporting Evidence: file:yeast/PDX3/PDX3-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion intermembrane space |
| GO:0005829 cytosol | RCA GO_REF:0000123 | MARK AS OVER ANNOTATED | Summary: YeastPathways default compartment for PLPSAL-PWY. No experimental evidence places Pdx3 in the cytosol; the only experimental localization is the mitochondrial intermembrane space. A cytosolic pool is possible (IMS proteins with no presequence often are dual) but unproven. Reason: The only basis for cytosol is the YeastPathways default compartment for the salvage pathway (RCA), not a localization assay. The one experimental localization of Pdx3 is the mitochondrial intermembrane space, and the IMS proteome paper notes that dual IMS/cytosol localization has been demonstrated only for Sod1 among its proteins. A cytosolic pool remains possible and is raised as a suggested question, but a pathway-default location is an over-annotation until shown. Decided rather than left UNDECIDED because the row's source is identified and carries no localization evidence. Supporting Evidence: PMID:22984289 So far only the superoxide dismutase Sod1 has been dually localized in the IMS and the cytosol PMID:22984289 YBR035CPDX3Pyridoxine phosphate oxidase file:yeast/PDX3/PDX3-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion intermembrane space |
| GO:0008615 pyridoxine biosynthetic process | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO from IPR000659. In bacteria PdxH is the final step of DXP-dependent de novo PLP synthesis, but Pdx3 neither makes pyridoxine nor acts in a de novo pathway; yeast de novo PLP is made by Snz1/Sno1. Pdx3 converts PNP/PMP to PLP in salvage. Reason: Wrong process for a yeast PNPOx; replace with PLP salvage. Proposed replacements: pyridoxal 5'-phosphate salvage Supporting Evidence: file:yeast/PDX3/PDX3-uniprot.txt PATHWAY: Cofactor metabolism; pyridoxal 5'-phosphate salvage; pyridoxal |
| GO:0009443 pyridoxal 5'-phosphate salvage | IC PMID:7896706 Sterol uptake induced by an impairment of pyridoxal phosphat... | ACCEPT | Summary: IC from the oxidase activity (Loubbardi 1995). pdx3 mutants cannot use pyridoxine but can use pyridoxal or pyridoxamine, matching a salvage oxidase role. Reason: Pdx3 itself catalyses the PNP/PMP oxidation step of B6 salvage (yeast makes PLP de novo through Snz1/Sno1), and pdx3 mutants cannot use pyridoxine. Supporting Evidence: PMID:7896706 These mutants are characterized by a lack in pyridoxine (pyridoxamine) phosphate oxidase [P(N/M)P oxidase] (EC 1.4.3.5) activity. |
| GO:0009443 pyridoxal 5'-phosphate salvage | RCA GO_REF:0000123 | ACCEPT | Summary: YeastPathways PLPSAL-PWY. Correct process for Pdx3. Reason: Consistent with the IC and the UniProt salvage pathway lines; correct process. Supporting Evidence: file:yeast/PDX3/PDX3-uniprot.txt PATHWAY: Cofactor metabolism; pyridoxal 5'-phosphate salvage; pyridoxal |
| GO:0010181 FMN binding | IDA PMID:26327315 Experimental Evidence for a Revision in the Annotation of Pu... | ACCEPT | Summary: IDA from Domitrovic et al. 2015; purified Pdx3 is yellow and quenches FMN fluorescence, and the crystal structure contains bound FMN. FMN is the catalytic cofactor. Reason: FMN is the catalytic prosthetic group of PNPOx (hydride acceptor); binding is integral to the activity. Accepted, as for E. coli and P. putida PdxH and human PNPO. Supporting Evidence: PMID:26327315 Note that purified Pdx3 presents the characteristic yellow color of proteins that are bound to FMN. |
| GO:0010181 FMN binding | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO from IPR000659; correct cofactor binding for this flavoenzyme. Reason: Same claim as the IDA row; the IPR000659 signature fits a full two-domain PNPOx. Supporting Evidence: PMID:26327315 Note that purified Pdx3 presents the characteristic yellow color of proteins that are bound to FMN. |
| GO:0016638 oxidoreductase activity, acting on the CH-NH2 group of donors | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO from the PNPOx conserved-site signature IPR019740. Correct (PMP oxidation acts on a CH-NH2 donor) but a general parent of pyridoxamine phosphate oxidase activity. Reason: Generic parent term; the specific activity is established. Proposed replacements: pyridoxamine phosphate oxidase activity Supporting Evidence: PMID:26327315 On the other hand, Pdx3 recombinant protein exhibits all expected properties of a bona fide P(N/M)P oxidase. |
| GO:0042816 vitamin B6 metabolic process | IEA GO_REF:0000117 | MODIFY | Summary: ARBA rule giving the general vitamin B6 metabolic process; an ancestor of PLP salvage, which is the specific process for Pdx3. Reason: Too general. Proposed replacements: pyridoxal 5'-phosphate salvage Supporting Evidence: file:yeast/PDX3/PDX3-uniprot.txt PATHWAY: Cofactor metabolism; pyridoxal 5'-phosphate salvage; pyridoxal |
| GO:0042823 pyridoxal 5'-phosphate biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN000091366 (E. coli PdxH, M. tuberculosis, human, fly, Arabidopsis). PLP biosynthetic process is the parent of PLP salvage and correct for Pdx3, whose product is PLP. Reason: PTN000091366 places PLP biosynthesis at the family root. For Pdx3 the general term is correct (its product is PLP) and is kept as the family-level process claim; the yeast route is recorded separately by the accepted GO:0009443 salvage rows. Supporting Evidence: file:interpro/panther/PTHR10851/PTHR10851-paint.tsv PTHR10851 PTN000091366 GO:0042823 P IBD false PMID:26327315 On the other hand, Pdx3 recombinant protein exhibits all expected properties of a bona fide P(N/M)P oxidase. |
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Download this section (compressed HTML)Q: Is Pdx3 exclusively in the mitochondrial intermembrane space, or is there a cytosolic pool, and how is PLP made in the IMS exported to cytosolic PLP-dependent enzymes?
Experiment: Determine Pdx3 sub-cellular distribution by fractionation and protease protection with tagged and untagged protein, and test whether a cytosol-targeted Pdx3 rescues pyridoxine utilisation in pdx3 cells.
Pyridoxine (pyridoxamine) phosphate oxidase; has homologs in E. coli and Myxococcus xanthus; transcription is under the general control of nitrogen metabolism
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