PNPO (human) — gene review notes

Identity

• UniProt: Q9NVS9 (PNPO_HUMAN), 261 aa, chromosome 17. HGNC:30260.
• RecName: Pyridoxine-5'-phosphate oxidase; AltName: Pyridoxamine-phosphate oxidase. EC 1.4.3.5.
  [file:human/PNPO/PNPO-uniprot.txt "RecName: Full=Pyridoxine-5'-phosphate oxidase"]
  [file:human/PNPO/PNPO-uniprot.txt "AltName: Full=Pyridoxamine-phosphate oxidase"]

Core molecular function — FMN-dependent oxidase

• Catalyzes the terminal, rate-limiting step of pyridoxal 5'-phosphate (PLP, active vitamin B6) synthesis:
  oxidation of pyridoxine 5'-phosphate (PNP) OR pyridoxamine 5'-phosphate (PMP) to PLP.
  [file:human/PNPO/PNPO-uniprot.txt "Catalyzes the oxidation of either pyridoxine 5'-phosphate"]
• Reaction 1: PNP + O2 = PLP + H2O2 (Rhea:15149; EC 1.4.3.5).
  [file:human/PNPO/PNPO-uniprot.txt "pyridoxine 5'-phosphate + O2 = pyridoxal 5'-phosphate + H2O2"]
• Reaction 2: PMP + O2 + H2O = PLP + H2O2 + NH4(+) (Rhea:15817).
  [file:human/PNPO/PNPO-uniprot.txt "pyridoxamine 5'-phosphate + O2 + H2O = pyridoxal 5'-phosphate"]
• Cofactor: FMN, 1 per subunit (flavoprotein).
  [file:human/PNPO/PNPO-uniprot.txt "Binds 1 FMN per subunit"]
• GO:0004733 "pyridoxamine phosphate oxidase activity" is the correct catalytic MF; the GO definition
  explicitly covers BOTH PNP and PMP oxidation ("This activity can also oxidize pyridoxine 5'-phosphate
  to pyridoxal 5'-phosphate + H2O2") — verified via OLS. So the single GO term captures both UniProt reactions.
• Kinetics: KM 2.1 uM (PNP), 6.2 uM (PMP); low turnover (~0.2 s^-1).
  [file:human/PNPO/PNPO-uniprot.txt "KM=2.1 uM for pyridoxine 5'-phosphate"]
  PMID:12824491

Structure / quaternary

• Homodimer; binds one PLP tightly per subunit (product inhibition).
  [file:human/PNPO/PNPO-uniprot.txt "SUBUNIT: Homodimer"]
  PMID:12824491
  PMID:12824491
• Crystal structure 1.95 A of residues 49-161 in complex with FMN and PLP (PDB 1NRG), plus later
  structures (6H00, 8ROS, etc.). Fold = FMN-binding split barrel; PNPOx family.
  [file:human/PNPO/PNPO-uniprot.txt "SIMILARITY: Belongs to the pyridoxamine 5'-phosphate oxidase family"]
• Product inhibition by PLP: "Pyridoxal 5'-phosphate is an effective product inhibitor."
  PMID:12824491

Domain / truncation biology (PMID:15182361)

• Human brain PNPO expressed in E. coli; N- and C-terminal truncation series.
• Deletion of N-terminal 56 residues does NOT affect coenzyme binding or catalysis (so the N-terminal
  ~56 aa is dispensable), whereas deletion of residues 1-72 or the C-terminal 238-261 abolishes activity.
  PMID:15182361
  [file:human/PNPO/PNPO-uniprot.txt "1..56 ... Missing: Has no effect on the catalytic activity"]
  [file:human/PNPO/PNPO-uniprot.txt "238..261 ... Missing: Loss of catalytic activity"]
• Tissue distribution: ubiquitous ("housekeeping"), highest in liver, skeletal muscle, kidney.
  PMID:15182361

Pathway / localization

• Pathway: cofactor metabolism; PLP salvage — PLP from PMP (step 1/1) and PLP from PNP (step 1/1).
  [file:human/PNPO/PNPO-uniprot.txt "Cofactor metabolism; pyridoxal 5'-phosphate salvage"]
• Reactome: Vitamin B6 activation to pyridoxal phosphate (R-HSA-964975); reactions R-HSA-965019
  (PDXP->PXLP) and R-HSA-965079 (PXAP->PXLP), both localized to cytosol.
  [file:human/PNPO/PNPO-uniprot.txt "Reactome; R-HSA-964975; Vitamin B6 activation to pyridoxal phosphate"]
• Localization: cytosol (Reactome TAS; consistent with a soluble metabolic enzyme). A mitochondrion
  call exists via IBA (GO_Central) and HTP (FlyBase, PMID:34800366) but PNPO is canonically cytosolic;
  the cached PMID:34800366 (mouse mitochondrial proteome, Cell Metab 2021) does not mention PNPO in its
  extracted text. Treated as over-annotation, not removed (HTP is experimental-derived; IBA propagated).

Disease

• PNPO deficiency (PNPOD, MIM:610090): neonatal/early-infantile epileptic encephalopathy; seizures
  respond to PLP but are resistant to pyridoxine.
  [file:human/PNPO/PNPO-uniprot.txt "onset within hours of birth of a severe seizure disorder"]
  PMID:15772097
• Pathogenic variants R229W (strong activity decrease), R229Q, R225H; expression studies confirmed
  R229W markedly reduces oxidase activity, splice/stop mutations null.
  PMID:15772097
  [file:human/PNPO/PNPO-uniprot.txt "R -> W (in PNPOD; strong activity decrease"]

Interactions (GOA GO:0005515 IPI)

• Six bare "protein binding" IPI annotations from proteome-scale interactome / IntAct screens
  (PMID:25910212, 28514442, 32296183, 33961781) with partners AGTRAP(Q6RW13-2), CMTM5(Q96DZ9-2),
  LIME1(Q9H400), MTERF1(Q99551). UniProt lists the same IntAct partners.
  [file:human/PNPO/PNPO-uniprot.txt "AGTRAP; NbExp=6"]
• These are non-informative "protein binding" (GO:0005515) with no functional/complex interpretation for
  a metabolic enzyme; kept but MARK_AS_OVER_ANNOTATED per curation policy (do not REMOVE IPI).

Review decisions summary

• Core MF: GO:0004733 (pyridoxamine phosphate oxidase activity, covers PNP+PMP) + GO:0010181 (FMN binding).
• Core BP: GO:0042823 (PLP biosynthetic process). GO:0008615 (pyridoxine biosynthetic process, IEA-KW) is
  a mislabel of the pathway (PNPO makes PLP from already-phosphorylated B6 vitamers, it does not synthesize
  pyridoxine) -> MODIFY to GO:0042823 / KEEP as vitamin B6 biosynthesis context.
• Cofactor-binding MFs GO:0030170 (PLP binding) and GO:0042803 (homodimerization) are real (structure)
  but supportive, not the core catalytic function -> ACCEPT / KEEP_AS_NON_CORE.
• Location cytosol (Reactome TAS) ACCEPT; mitochondrion (IBA + HTP) MARK_AS_OVER_ANNOTATED.

2026-11-03 ORGANELLES batch 4 (mitochondrion rows)

Rows: GO:0005739 IBA (GO_REF:0000033, is_active_in) and GO:0005739 HTP (PMID:34800366,
MitoCoP, located_in). No sub-compartment rows.

Checks:
- PAINT: interpro/panther/PTHR10851/PTHR10851-paint.tsv puts GO:0005739 at PTN000091367
(taxon:2759), seeds AGI_LocusCode:AT5G49970 (PPOX1/PDX3, Q9LTX3) and SGD:S000000239 (PDX3,
P38075), identities checked via UniProt REST. QuickGO (2026-11-03): Q9LTX3 has TAIR IDA
mitochondrion (PMID:24599492); P38075 has SGD IDA GO:0005758 intermembrane space
(PMID:22984289). The node is soundly seeded.
- MitoCoP (PMID:34800366) is a human proteome, not mouse as the earlier notes said
PMID:34800366.
Per-protein calls are in supplementary tables, not in the cached text.
- MitoCarta3.0: PNPO listed; evidence "APEX_matrix, yeast mito homolog++, coexpression, MS/MS";
SubMitoLocalization "Matrix"; MSMS "0-25pure" (2 mouse tissues); TargetP score 5; HPA column
"Cytosol;Nucleoplasm (Enhanced)". Not a cached, citable source.
- UniProt Q9NVS9 has no subcellular-location comment, no transit peptide, no TM. Reactome puts
both reactions in the cytosol.

Outcome: both rows MARK_AS_OVER_ANNOTATED -> KEEP_AS_NON_CORE (DTYMK precedent: MitoCoP plus
MitoCarta3.0 with an in-cell APEX line; bulk enzyme cytosolic). IBA propagation_review root
cause PROPAGATION_BAD -> NO_FAILURE_NON_CORE. PMID:34800366 reference_review changed from
MISCITED to VERIFIED. is_active_in: not shown (no PNPO activity measured in human mitochondria).
Stay at the root: the sub-compartment lines disagree (APEX matrix vs yeast IMS seed) and
there is no targeting signal.