PPOX

Existing Annotations Review

GO Term	Evidence	Action	Reason
GO:0004729 protoporphyrinogen oxidase activity, oxygen as acceptor	IBA GO_REF:0000033	ACCEPT	Summary: Phylogenetic (IBA) assignment of the defining catalytic activity of PPOX, transferred across the protoporphyrinogen oxidase orthology group. This is the core molecular function and is strongly supported by direct human enzyme assays. Reason: PAINT IBA (GO_REF:0000033) within PANTHER family PTHR42923 (protoporphyrinogen oxidase). Oxygen-dependent protoporphyrinogen oxidase (EC 1.3.3.4) is the experimentally established activity of the human enzyme (assays, crystal structure, kinetics). The current GO label for this id is 'oxygen-dependent protoporphyrinogen oxidase activity'. Supporting Evidence: PMID:7713909 Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in the penultimate step in the heme biosynthetic pathway. file:human/PPOX/PPOX-uniprot.txt Reaction=protoporphyrinogen IX + 3 O2 = protoporphyrin IX + 3 H2O2 file:human/PPOX/PPOX-uniprot.txt PANTHER; PTHR42923; PROTOPORPHYRINOGEN OXIDASE; 1.
GO:0006783 heme biosynthetic process	IBA GO_REF:0000033	ACCEPT	Summary: Phylogenetic (IBA) assignment placing PPOX in the heme biosynthetic pathway. This is the core biological process for the gene: the PPOX-catalysed step is the penultimate reaction of heme synthesis. Reason: PAINT IBA (GO_REF:0000033) from PTHR42923. PPOX itself performs the seventh step of heme biosynthesis; deficiency causes variegate porphyria. Core process. Supporting Evidence: PMID:7713909 Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in the penultimate step in the heme biosynthetic pathway. PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway. file:human/PPOX/PPOX-uniprot.txt PANTHER; PTHR42923; PROTOPORPHYRINOGEN OXIDASE; 1.
GO:0005743 mitochondrial inner membrane	IBA GO_REF:0000033	ACCEPT	Summary: Phylogenetic (IBA) assignment of PPOX to the mitochondrial inner membrane, its established site of action. PPOX is a peripheral membrane protein on the intermembrane-space-facing (outer) surface of the inner membrane. Reason: The human enzyme is a mitochondrial inner-membrane protein; the crystallographic study explicitly describes hPPO as a mitochondrial inner membrane protein, and Reactome places it on the outer surface of the inner mitochondrial membrane. Core cellular location. Supporting Evidence: PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway. Reactome:R-HSA-189423 The protein resides on the outer surface of the inner mitochondrial membrane. file:human/PPOX/PPOX-uniprot.txt PANTHER; PTHR42923; PROTOPORPHYRINOGEN OXIDASE; 1.
GO:0004729 protoporphyrinogen oxidase activity, oxygen as acceptor	IEA GO_REF:0000120	ACCEPT	Summary: Electronic assignment (combined IEA methods, tied to EC 1.3.3.4, RHEA:25576 and InterPro:IPR004572) of the core catalytic activity. Redundant with the experimental and IBA annotations to the same term. Reason: Correct core molecular function, independently supported by direct enzyme assays and the mapped EC number/RHEA reaction. Duplicate ID with different evidence is acceptable. Supporting Evidence: file:human/PPOX/PPOX-uniprot.txt Reaction=protoporphyrinogen IX + 3 O2 = protoporphyrin IX + 3 H2O2 file:human/PPOX/PPOX-uniprot.txt FUNCTION: Catalyzes the 6-electron oxidation of protoporphyrinogen-IX to form protoporphyrin-IX. file:human/PPOX/PPOX-uniprot.txt InterPro; IPR004572; Protoporphyrinogen_oxidase.
GO:0005743 mitochondrial inner membrane	IEA GO_REF:0000120	ACCEPT	Summary: Electronic (combined IEA / SubCell SL-0168) assignment of mitochondrial inner membrane localisation, matching the experimentally supported location. Reason: Consistent with the UniProt subcellular location (mitochondrion inner membrane, peripheral, intermembrane side) and with the crystallographic characterisation of the human enzyme. Core location. Supporting Evidence: file:human/PPOX/PPOX-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion inner membrane PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway.
GO:0005758 mitochondrial intermembrane space	IEA GO_REF:0000117	MARK AS OVER ANNOTATED	Summary: ARBA electronic assignment to the mitochondrial intermembrane space. PPOX faces the intermembrane space (it is a peripheral protein on the outer/IMS-facing surface of the inner membrane) but is not a soluble intermembrane-space protein. Reason: PPOX is a peripheral protein anchored to the inner membrane with its active site facing the intermembrane space; UniProt records it as inner membrane, peripheral, intermembrane side, and Reactome places it on the outer surface of the inner membrane. Calling it an intermembrane-space protein overstates a free luminal location; the accurate component is mitochondrial inner membrane (GO:0005743), already carried. Marked as over-annotated rather than removed because the protein does face, and act on substrate in, the intermembrane space. Supporting Evidence: file:human/PPOX/PPOX-uniprot.txt Peripheral membrane protein {ECO:0000250\|UniProtKB:P51175}; Intermembrane side Reactome:R-HSA-189423 The protein resides on the outer surface of the inner mitochondrial membrane.
GO:0006779 porphyrin-containing compound biosynthetic process	IEA GO_REF:0000120	MODIFY	Summary: Electronic assignment to porphyrin-containing compound biosynthesis. This is a correct but more general parent of the specific heme biosynthetic process; PPOX produces the porphyrin protoporphyrin IX. Reason: Correct but more general than the evidence supports. PPOX's product, protoporphyrin IX, is the direct substrate of ferrochelatase and is committed to heme in human cells, so the specific descendant heme biosynthetic process (GO:0006783), already carried by the gene, is the appropriate replacement. Proposed replacements: heme biosynthetic process Supporting Evidence: PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway. file:human/PPOX/PPOX-uniprot.txt FUNCTION: Catalyzes the 6-electron oxidation of protoporphyrinogen-IX to form protoporphyrin-IX.
GO:0006785 heme B biosynthetic process	IEA GO_REF:0000117	ACCEPT	Summary: ARBA electronic assignment to heme B (protoheme) biosynthesis, the specific branch of heme synthesis to which the PPOX-generated protoporphyrin IX is committed (protoporphyrin IX + Fe2+ -> heme B). Reason: Heme b (protoheme IX) is the immediate downstream product of the protoporphyrin IX made by PPOX (via ferrochelatase). The term is a valid, more specific descendant of heme biosynthetic process. Keep as a specific process; the general heme biosynthetic process remains the core BP. Supporting Evidence: PMID:7713909 Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in the penultimate step in the heme biosynthetic pathway. PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway.
GO:0016491 oxidoreductase activity	IEA GO_REF:0000002	MODIFY	Summary: InterPro2GO electronic annotation (GO_REF:0000002) to generic oxidoreductase activity. Reason: Correct but general. The specific activity is oxygen-dependent protoporphyrinogen oxidase activity (GO:0004729), demonstrated directly for the human enzyme and already carried by the gene; that is the appropriate replacement. Proposed replacements: oxygen-dependent protoporphyrinogen oxidase activity Supporting Evidence: file:human/PPOX/PPOX-uniprot.txt Reaction=protoporphyrinogen IX + 3 O2 = protoporphyrin IX + 3 H2O2 file:human/PPOX/PPOX-uniprot.txt InterPro; IPR004572; Protoporphyrinogen_oxidase.
GO:0031966 mitochondrial membrane	IEA GO_REF:0000117	MODIFY	Summary: ARBA electronic assignment to the generic mitochondrial membrane. Correct but broader than the specific mitochondrial inner membrane localisation. Reason: Correct but general. PPOX is specifically a peripheral protein of the mitochondrial inner membrane (crystal-structure study, UniProt, Reactome), so the specific descendant mitochondrial inner membrane (GO:0005743), already carried by the gene, is the appropriate replacement. Proposed replacements: mitochondrial inner membrane Supporting Evidence: file:human/PPOX/PPOX-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion inner membrane PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway.
GO:0006783 heme biosynthetic process	IEA GO_REF:0000107	ACCEPT	Summary: Ensembl (orthology-based) electronic transfer of the core heme biosynthetic process annotation. Redundant with the IBA/ISS/IDA annotations to the same term. Reason: Correct core biological process, supported by direct experimental evidence for the human enzyme; the electronic transfer from the rat ortholog is appropriate. Supporting Evidence: PMID:7713909 Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in the penultimate step in the heme biosynthetic pathway. file:human/PPOX/PPOX-uniprot.txt FUNCTION: Catalyzes the 6-electron oxidation of protoporphyrinogen-IX to form protoporphyrin-IX.
GO:0009410 response to xenobiotic stimulus	IEA GO_REF:0000107	KEEP AS NON CORE	Summary: Ensembl electronic transfer (from the rat ortholog D3ZVN7) of "response to xenobiotic stimulus". PPOX is the molecular target of diphenyl-ether herbicides (e.g. acifluorfen), which underlies this annotation, but responding to a xenobiotic is not a core evolved function of the enzyme. Reason: Ensembl Compara projection (GO_REF:0000107) from rat. The response definition covers a change in the activity of a gene product after a stimulus, and PPOX activity is specifically inhibited by xenobiotic diphenyl-ether herbicides such as acifluorfen (shown on the human enzyme). Kept as non-core pharmacology/toxicology biology, not a detoxification role. Supporting Evidence: PMID:7713909 A homogenate of the monkey kidney COS-1 cells that had been transfected with the cDNA had much higher PPO activity than an extract of control cells, and this activity was inhibited by acifluorfen, a specific inhibitor of PPO. PMID:21048046 we report the crystal structure of hPPO in complex with the coenzyme flavin adenine dinucleotide (FAD) and the inhibitor acifluorfen
GO:0004729 protoporphyrinogen oxidase activity, oxygen as acceptor	ISS GO_REF:0000024	ACCEPT	Summary: Curator sequence-similarity (ISS) transfer of the core catalytic activity from the mouse ortholog (MGI:104968). Redundant with the direct experimental and IBA annotations. Reason: Correct core molecular function; sequence-similarity transfer is fully consistent with the directly assayed human enzyme activity. Supporting Evidence: PMID:7713909 Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in the penultimate step in the heme biosynthetic pathway. file:human/PPOX/PPOX-uniprot.txt Reaction=protoporphyrinogen IX + 3 O2 = protoporphyrin IX + 3 H2O2
GO:0006783 heme biosynthetic process	ISS GO_REF:0000024	ACCEPT	Summary: Curator sequence-similarity (ISS) transfer of the core heme biosynthetic process from the mouse ortholog. Redundant with IBA/IEA/IDA annotations to the same term. Reason: Correct core biological process, consistent with all other lines of evidence. Supporting Evidence: PMID:7713909 Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in the penultimate step in the heme biosynthetic pathway.
GO:0004729 protoporphyrinogen oxidase activity, oxygen as acceptor	EXP PMID:23467411 Quantitative structural insight into human variegate porphyr...	ACCEPT	Summary: Experimental (EXP) annotation of protoporphyrinogen oxidase activity from the quantitative structural/kinetic study of wild-type and VP-mutant human PPO, which assayed kcat/Km of the enzyme and characterised the FAD- and O2-dependent oxidation of protoporphyrinogen IX to protoporphyrin IX. Reason: Direct experimental determination of the core catalytic activity of the human enzyme, including quantitative kinetics; the defining function of the gene. Supporting Evidence: PMID:23467411 catalyzes the oxidation of protoporphyrinogen IX (protogen) to protoporphyrin IX (porphyrin) in the presence of cofactor FAD and molecular oxygen
GO:0005743 mitochondrial inner membrane	ISS GO_REF:0000024	ACCEPT	Summary: Curator sequence-similarity (ISS) transfer of mitochondrial inner membrane localisation from the mouse ortholog (P51175). Consistent with the experimentally supported location. Reason: Correct core location; matches the UniProt subcellular location and the crystallographic description of the human enzyme. Supporting Evidence: PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway. file:human/PPOX/PPOX-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion inner membrane
GO:0005743 mitochondrial inner membrane	ISS GO_REF:0000024	ACCEPT	Summary: Duplicate GOA line of the curator sequence-similarity (ISS) transfer of mitochondrial inner membrane localisation from the mouse ortholog (P51175); differs only in annotation date/source. Same conclusion as the other ISS localisation annotation. Reason: Correct core location; matches the UniProt subcellular location and the crystallographic description of the human enzyme. Duplicate of the equivalent GO:0005743 ISS annotation. Supporting Evidence: PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway. file:human/PPOX/PPOX-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion inner membrane
GO:0004729 protoporphyrinogen oxidase activity, oxygen as acceptor	IDA PMID:21048046 Structural insight into human variegate porphyria disease.	ACCEPT	Summary: Direct assay (IDA) of protoporphyrinogen oxidase activity from the 1.9 Angstrom crystal structure study of human PPO in complex with FAD and the inhibitor acifluorfen, which biochemically characterised the wild-type enzyme and 47 VP-causing mutants. Reason: Direct experimental evidence for the core catalytic function of the human enzyme, including structural and kinetic characterisation. Defining molecular function. Supporting Evidence: PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway. PMID:21048046 we report the crystal structure of hPPO in complex with the coenzyme flavin adenine dinucleotide (FAD) and the inhibitor acifluorfen
GO:0006785 heme B biosynthetic process	IDA PMID:7713909 Cloning of a human cDNA for protoporphyrinogen oxidase by co...	ACCEPT	Summary: Direct assay (IDA) placing PPOX in heme (heme B / protoheme) biosynthesis, from the cDNA cloning study that complemented an E. coli hemG (PPO-deficient) mutant and demonstrated acifluorfen-inhibitable PPO activity in transfected cells. Reason: Functional complementation of a heme-pathway (hemG) mutant plus demonstration of PPO activity directly establishes PPOX's role in producing the protoporphyrin IX committed to heme b synthesis. Keep as a specific heme-branch process alongside the general core heme biosynthetic process. Supporting Evidence: PMID:7713909 Thus, a hemG mutant strain of Escherichia coli deficient in PPO is defective in aerobic respiration and grows poorly even in rich medium. PMID:7713909 By complementation with a human placental cDNA library, we were able to isolate a clone that enhanced the poor growth of such a hemG mutant strain.
GO:0005739 mitochondrion	HTP PMID:34800366 Quantitative high-confidence human mitochondrial proteome an...	ACCEPT	Summary: High-throughput (HTP) proteomic assignment of PPOX to the mitochondrion, from a quantitative high-confidence human mitochondrial proteome (MitoCoP) study. Broader than, but consistent with, the specific inner-membrane localisation. Reason: Direct high-throughput detection in the human MitoCoP mitochondrial proteome. This organelle-level assay cannot resolve the sub-compartment, so per the root-organelle policy the row is accepted at the organelle; the inner-membrane location is carried by separate rows. Supporting Evidence: PMID:34800366 defined a mitochondrial high-confidence proteome of >1,100 proteins (MitoCoP)
GO:0005758 mitochondrial intermembrane space	TAS Reactome:R-HSA-189423	MARK AS OVER ANNOTATED	Summary: Reactome traceable-author-statement placing PPO on the outer surface of the inner mitochondrial membrane (facing the intermembrane space). As with the IEA IMS annotation, the intermembrane-space term overstates a soluble-luminal localisation. Reason: PPOX is a peripheral protein anchored to the inner membrane with its active site facing the intermembrane space; UniProt records it as inner membrane, peripheral, intermembrane side, and Reactome places it on the outer surface of the inner membrane. Calling it an intermembrane-space protein overstates a free luminal location; the accurate component is mitochondrial inner membrane (GO:0005743), already carried. Marked as over-annotated rather than removed because the protein does face, and act on substrate in, the intermembrane space. Supporting Evidence: Reactome:R-HSA-189423 The protein resides on the outer surface of the inner mitochondrial membrane. file:human/PPOX/PPOX-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion inner membrane
GO:0004729 protoporphyrinogen oxidase activity, oxygen as acceptor	IDA PMID:7713909 Cloning of a human cDNA for protoporphyrinogen oxidase by co...	ACCEPT	Summary: Direct assay (IDA) of protoporphyrinogen oxidase activity from the original human cDNA cloning: COS-1 cells transfected with the cDNA showed markedly higher, acifluorfen-inhibitable PPO activity than controls. Reason: Direct experimental demonstration of the core catalytic activity of the human gene product. Defining molecular function. Supporting Evidence: PMID:7713909 A homogenate of the monkey kidney COS-1 cells that had been transfected with the cDNA had much higher PPO activity than an extract of control cells, and this activity was inhibited by acifluorfen, a specific inhibitor of PPO.
GO:0006779 porphyrin-containing compound biosynthetic process	IDA PMID:7713909 Cloning of a human cDNA for protoporphyrinogen oxidase by co...	MODIFY	Summary: Direct assay (IDA) placing PPOX in porphyrin-containing compound biosynthesis, from the cloning/complementation study. PPOX generates the porphyrin protoporphyrin IX. Reason: Correct but more general than the evidence supports. PPOX's product, protoporphyrin IX, is the direct substrate of ferrochelatase and is committed to heme in human cells, so the specific descendant heme biosynthetic process (GO:0006783), already carried by the gene, is the appropriate replacement. Proposed replacements: heme biosynthetic process Supporting Evidence: PMID:7713909 Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in the penultimate step in the heme biosynthetic pathway.
GO:0006783 heme biosynthetic process	IDA PMID:7713909 Cloning of a human cDNA for protoporphyrinogen oxidase by co...	ACCEPT	Summary: Direct assay (IDA) of PPOX's role in heme biosynthesis, from functional complementation of an E. coli hemG (PPO-deficient) mutant with the human cDNA. This is the core biological process for the gene. Reason: Functional complementation of a heme-pathway mutant and restoration of PPO activity directly establishes the role of PPOX in heme biosynthesis. Core BP. Supporting Evidence: PMID:7713909 Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in the penultimate step in the heme biosynthetic pathway. PMID:7713909 By complementation with a human placental cDNA library, we were able to isolate a clone that enhanced the poor growth of such a hemG mutant strain.
GO:0031966 mitochondrial membrane	IMP PMID:7713909 Cloning of a human cDNA for protoporphyrinogen oxidase by co...	MODIFY	Summary: Mutant-phenotype-based (IMP) localisation to the mitochondrial membrane, from the cloning study showing the in vitro-translated protein associates with isolated mitochondria without size change, indicating no cleaved targeting presequence. Reason: Correct but general. PPOX is specifically a peripheral protein of the mitochondrial inner membrane (crystal-structure study, UniProt, Reactome), so the specific descendant mitochondrial inner membrane (GO:0005743), already carried by the gene, is the appropriate replacement. The cited import experiment supports mitochondrial association but not the sub-membrane. Proposed replacements: mitochondrial inner membrane Supporting Evidence: PMID:7713909 after incubation with isolated mitochondria the protein was found to be located in the mitochondria, having just the same size as before, an indication that PPO is a mitochondrial enzyme and has no apparent transport-specific leader sequence. PMID:21048046 Human protoporphyrinogen IX oxidase (hPPO), a mitochondrial inner membrane protein, converts protoporphyrinogen IX to protoporphyrin IX in the heme biosynthetic pathway.
GO:0050660 flavin adenine dinucleotide binding	TAS PMID:7713909 Cloning of a human cDNA for protoporphyrinogen oxidase by co...	ACCEPT	Summary: Traceable author statement that PPOX binds FAD. The N-terminal sequence contains a classic dinucleotide-binding motif, and the crystal structure resolves one FAD bound per subunit; FAD is the essential redox cofactor for the oxidation reaction. Reason: PPOX is a flavoprotein that binds one FAD per subunit as its catalytic cofactor, confirmed both by sequence motif and by crystal structures. This is a genuine, though secondary/enabling, molecular function supporting the core oxidase activity. Supporting Evidence: PMID:7713909 The NH2-terminal amino acid sequence of the deduced PPO contains a conserved amino acid sequence that forms the dinucleotide-binding site in many flavin-containing proteins. file:human/PPOX/PPOX-uniprot.txt Note=Binds 1 FAD per subunit.

Gene Description

Existing Annotations Review

Core Functions

FAD- and oxygen-dependent six-electron oxidation of protoporphyrinogen IX to protoporphyrin IX, the seventh (penultimate) step of heme biosynthesis, catalysed at the mitochondrial inner membrane.

Binds one non-covalent FAD cofactor per subunit, the flavin redox centre required for the oxidase activity.

References

📚 Additional Documentation

Notes

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