PA1204 is a small (185 aa) cytoplasmic flavoprotein of the SsuE/chromate reductase (FMN_red, PF03358) family of Pseudomonas aeruginosa. Its 1.28 A crystal structure shows a flavodoxin-like homodimer with a non-canonical FMN-binding motif (GSLRSGSYN), and the purified protein binds FMN and shows NAD(P)H-dependent FMN reductase activity. It also reduces quinones by a two-electron route with little semiquinone formation, placing it with the soluble NAD(P)H:quinone oxidoreductases (E. coli ChrR, Synechocystis ArsH, yeast Lot6) that are thought to protect against redox-cycling quinones; overexpression of the P. aeruginosa enzymes in this group gave no clear H2O2-protective phenotype except for the WrbA and MdaB orthologs. PA1204 lies in the pa1202-pa1205 operon that is strongly derepressed in bexR mutants.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003955 NAD(P)H dehydrogenase (quinone) activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from the PTN001254275 IBD (4 seeds: yeast Lot6, E. coli ChrR P0AGE6, Synechocystis ArsH P74312 and PA1204 itself). PA1204 is a seed, which is expected, not circular. Reason: PA1204 reduces quinones in vitro by a two-electron route, and the other three seeds are characterised NAD(P)H:quinone reductases, so the IBD stands with or without this seed. Core MF. Supporting Evidence: PMID:25085734 In in vitro assays, each enzyme was shown to reduce quinone substrates with only minimal semiquinone formation. PMID:24915188 Recent data have shown that PA0949, PA1204, PA2580 and PA4975, have NAD(P)H quinone oxidoreductase activities [75]. |
| GO:0003955 NAD(P)H dehydrogenase (quinone) activity | IMP PMID:25085734 Pseudomonas aeruginosa MdaB and WrbA are water-soluble two-e... | ACCEPT | Summary: PseudoCAP IMP and one of the four seeds of the PTN001254275 GO:0003955 IBD. The paper assayed purified P. aeruginosa quinone oxidoreductase orthologs in vitro. Seed verdict SOUND for the term; the evidence code should be IDA. Reason: The activity is supported by the in vitro assays in the abstract and by Ryan et al. 2014, which names PA1204 among the enzymes with this activity. The evidence is a direct enzyme assay, not a mutant phenotype, so the IMP code is a source defect; the claim itself is accepted. Supporting Evidence: PMID:25085734 In in vitro assays, each enzyme was shown to reduce quinone substrates with only minimal semiquinone formation. |
| GO:0005829 cytosol | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: IBA cytosol from the same node (seeds Lot6 and ChrR). Reason: PA1204 is a soluble protein without signal peptide or transmembrane segment, and its quinone substrates and NAD(P)H are cytoplasmic; cytosol is the expected location. Location is peripheral to the function. Supporting Evidence: PMID:25085734 Water-soluble quinone oxidoreductases capable of reducing quinone substrates via a concerted two-electron mechanism have been implicated in bacterial antioxidant defence. |
| GO:0010181 FMN binding | IDA PMID:16552139 Structure determination of an FMN reductase from Pseudomonas... | ACCEPT | Summary: UniProt IDA from the crystal structure with bound FMN. Reason: FMN is the bound cofactor, located in the crystal structure. Supporting Evidence: PMID:16552139 it has been confirmed to bind flavin mononucleotide and the binding site was identified via X-ray crystallography |
| GO:0016491 oxidoreductase activity | IEA GO_REF:0000120 | TOO GENERAL | Summary: InterPro/ARBA mapping to oxidoreductase activity. Reason: True but uninformative; GO:0003955 NAD(P)H dehydrogenase (quinone) activity, accepted in this review, is_a descendant of GO:0016491. Supporting Evidence: PMID:16552139 NADPH-dependent activity has been confirmed with biochemical assays. |
| GO:0016655 oxidoreductase activity, acting on NAD(P)H, quinone or similar compound as acceptor | IDA PMID:24915188 Identification of NAD(P)H quinone oxidoreductase activity in... | TOO GENERAL | Summary: PseudoCAP IDA citing the Ryan et al. azoreductase paper, which mentions PA1204 quinone reductase activity only by citing other work. Reason: The claim is true (see GO:0003955), and GO:0003955, accepted in this review, is_a GO:0016655, so this parent is redundant. The cited paper did not assay PA1204; the IDA code is recorded as a source defect. Supporting Evidence: PMID:24915188 Recent data have shown that PA0949, PA1204, PA2580 and PA4975, have NAD(P)H quinone oxidoreductase activities [75]. |
| GO:0034599 cellular response to oxidative stress | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: IBA from the PTN001254275 IBD of GO:0034599, whose only seed is yeast Lot6. PA1204 is not a seed of this term. Reason: Two-electron quinone reductases are proposed to limit redox cycling, but in P. aeruginosa only the WrbA and MdaB orthologs gave an H2O2-protective phenotype when overexpressed in a katA mutant; no oxidative-stress phenotype has been reported for PA1204. The process is plausible for the family but not shown for this gene. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: FUNCTIONAL DIVERGENCE Sources checked: PANTHER:PTN001254275 · PTN001254275 UNRESOLVED Single-seed IBD (yeast Lot6) for a stress-response process across a 74-taxon clade of bacterial and fungal FMN reductases. Whether each member protects against oxidative stress depends on its physiological substrate, which is unknown for PA1204. SGD:S000004001 · LOT6 (yeast) SUPPORTS SOURCE BUT NOT TARGET Experimentally characterised yeast quinone reductase; its stress-response role is not shown for PA1204. Supporting Evidence: PMID:25085734 in a katA mutant, H2O2 challenged strains over-expressing the WrbA and MdaB orthologs grew significantly better than the empty plasmid control |
| GO:0042802 identical protein binding | IDA PMID:16552139 Structure determination of an FMN reductase from Pseudomonas... | TOO GENERAL | Summary: UniProt IDA; homodimer in the crystal structure. Reason: GO:0042803 protein homodimerization activity, kept in this review, is_a GO:0042802 and states the same observation more precisely. Supporting Evidence: file:PSEAE/PA1204/PA1204-uniprot.txt SUBUNIT: Homodimer. {ECO:0000269|PubMed:16552139}. |
| GO:0042803 protein homodimerization activity | IDA PMID:16552139 Structure determination of an FMN reductase from Pseudomonas... | KEEP AS NON CORE | Summary: UniProt IDA; homodimer in the crystal structure. Reason: The flavodoxin-like FMN reductases are dimeric; true but structural rather than the function. Supporting Evidence: file:PSEAE/PA1204/PA1204-uniprot.txt SUBUNIT: Homodimer. {ECO:0000269|PubMed:16552139}. |
| GO:0052873 FMN reductase (NADPH) activity | IDA PMID:16552139 Structure determination of an FMN reductase from Pseudomonas... | ACCEPT | Summary: UniProt IDA; NADPH-dependent FMN reductase activity of the purified protein. Reason: The structure paper confirmed NADPH-dependent activity biochemically, and UniProt records NAD(P)H-dependent FMN reductase activity from it. The physiological acceptor (free FMN versus quinones) is not settled, so both activities are kept as core. Supporting Evidence: PMID:16552139 NADPH-dependent activity has been confirmed with biochemical assays. file:PSEAE/PA1204/PA1204-uniprot.txt FUNCTION: Has NAD(P)H-dependent FMN reductase activity. |
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Download this section (compressed HTML)Q: Is the physiological electron acceptor of PA1204 a quinone (for example homogentisate quinone or phenazines), free FMN for a partner monooxygenase, or another substrate, and what does the BexR-controlled pa1202-pa1205 operon do?
Experiment: Steady-state kinetics of purified PA1204 with a panel of quinones, phenazines and free FMN, plus phenotyping of a PA1204 deletion (with and without katA) under quinone and H2O2 challenge.
Type: enzyme kinetics and mutant phenotyping
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