VanA (PP_3736) is the Rieske oxygenase subunit of the two-component VanAB vanillate O-demethylase in Pseudomonas putida KT2440. The protein carries an N-terminal Rieske [2Fe-2S] cluster-binding domain and a C-terminal VanA_C catalytic domain characteristic of aromatic-ring-hydroxylating oxygenase alpha subunits. The core function is oxygen-dependent oxidative O-demethylation of vanillate during lignin-derived aromatic catabolism; overexpression studies also show activity toward syringate, but vanillate O-demethylation remains the most specific native assignment for this gene.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003824 catalytic activity | IEA GO_REF:0000117 | MARK AS OVER ANNOTATED | Summary: VanA is certainly catalytic, but this term is far too broad to be informative. The same annotation set already contains the specific child term vanillate monooxygenase activity, which captures the actual biochemical role. Reason: GO:0003824 is a generic parent of the more specific enzyme activity already present. Retaining the parent term adds little value once GO:0018489 is accepted. |
| GO:0005506 iron ion binding | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: VanA is a Rieske oxygenase subunit with a [2Fe-2S] cluster. The general iron ion binding annotation reflects that cofactor requirement, but the specific cluster-binding term is already present and is more informative. Reason: GO:0005506 is true in a broad sense, but GO:0051537 precisely captures the relevant cofactor-binding chemistry for this protein and should be preferred. |
| GO:0016491 oxidoreductase activity | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: VanA is an oxidoreductase, but this parent term is less informative than the specific vanillate monooxygenase activity annotation supported by the UniProt EC assignment and the KT2440 vanAB literature. Reason: The annotation is redundant with GO:0018489, which states the actual substrate and reaction class. |
| GO:0018489 vanillate monooxygenase activity | IEA GO_REF:0000003 | ACCEPT | Summary: This is the core molecular function of VanA. UniProt assigns EC 1.14.13.82 to Q88GI6, recent KT2440 studies describe VanAB as the native Rieske non-heme iron monooxygenase used for vanillate O-demethylation, and vanAB engineering changes vanillate utilization in vivo. Reason: GO:0018489 is the most specific and biologically appropriate molecular function for vanA in KT2440. Falcon deep research confirms VanA is the terminal oxygenase of the two-component VanAB vanillate O-demethylase that converts vanillate to protocatechuate with release of formaldehyde. Supporting Evidence: file:PSEPK/vanA/vanA-uniprot.txt DE SubName: Full=Vanillate O-demethylase oxygenase subunit file:PSEPK/vanA/vanA-uniprot.txt DE EC=1.14.13.82 file:PSEPK/vanA/vanA-deep-research-falcon.md the two-component **VanAB** vanillate O-demethylase system that catalyzes oxidative demethylation of vanillate to protocatechuate, releasing **formaldehyde** as a coproduct file:PSEPK/vanA/vanA-deep-research-falcon.md VanA (Q88GI6; PP_3736)** is the **terminal oxygenase** subunit responsible for substrate hydroxylation/oxidative demethylation chemistry |
| GO:0051537 2 iron, 2 sulfur cluster binding | IEA GO_REF:0000002 | ACCEPT | Summary: VanA contains the canonical Rieske [2Fe-2S] domain and this cofactor is a defining feature of the oxygenase subunit. The annotation is specific and consistent with both the UniProt domain architecture and the literature description of VanAB as a Rieske monooxygenase. Reason: This is a specific, mechanistically relevant molecular function that explains how the oxygenase subunit supports catalytic turnover. Supporting Evidence: file:PSEPK/vanA/vanA-uniprot.txt DR GO; GO:0051537; F:2 iron, 2 sulfur cluster binding; IEA:UniProtKB-KW. file:PSEPK/vanA/vanA-uniprot.txt FT DOMAIN 7..107 file:PSEPK/vanA/vanA-deep-research-falcon.md the oxygenase (VanA-family) contains a **Rieske [2Fe–2S] cluster** and a **non-heme iron** catalytic center file:PSEPK/vanA/vanA-deep-research-falcon.md VanAB belongs to the **Rieske non-heme iron monooxygenase** |
| GO:0046191 aerobic phenol-containing compound catabolic process | IMP PMID:31809239 Biotransformation of corn bran derived ferulic acid to vanil... | NEW | Summary: Proposed new annotation. VanA participates in aerobic breakdown of methoxylated phenolic lignin-derived aromatics, most clearly vanillate. vanAB disruption blocks vanillic acid metabolism in KT2440, and recent KT2440 studies use native VanAB as the key O-demethylation step for vanillate and related substrates. Reason: The current GOA set lacks a biological process term capturing VanA's role in aerobic aromatic catabolism. GO:0046191 is broad but appropriate for the native vanillate/syringate O-demethylation context. Supporting Evidence: file:PSEPK/vanA/vanA-notes.md a vanAB nonfunctional mutant was explicitly selected because it was defective in vanillic acid metabolism file:PSEPK/vanA/vanA-deep-research-falcon.md VanAB enables growth on vanillate as a sole carbon/energy source by converting it to protocatechuate file:PSEPK/vanA/vanA-deep-research-falcon.md VanA operates in the **upper funneling pathway for lignin-derived guaiacyl aromatics** |
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Download this section (compressed HTML)Q: Is syringate oxidation by native KT2440 VanAB physiologically relevant at endogenous vanAB expression levels, or mainly an overexpression phenotype?
Q: Which native transcription factors, in addition to carbon catabolite repression, dominate vanAB induction during growth on vanillate in KT2440?
Q: Does vanA contribute measurably to substrate hierarchy when KT2440 is exposed to mixed lignin-derived aromatics?
Experiment: Construct clean vanA and vanB deletion/complementation strains and compare growth plus substrate disappearance on vanillate, vanillin-derived vanillate, and syringate under identical aerobic conditions.
Hypothesis: VanA is essential for native vanillate O-demethylation and any measurable syringate O-demethylation in KT2440.
Type: growth phenotype and substrate consumption assay
Experiment: Purify the VanAB complex and measure oxygen consumption plus product formation with vanillate, syringate, and additional methoxylated aromatics.
Hypothesis: VanA has highest catalytic efficiency on vanillate but retains measurable side activity on selected lignin-derived methoxylated phenolics.
Type: biochemical enzyme assay
Experiment: Perform RNA-seq or targeted promoter-reporter assays during growth on vanillate versus mixed aromatic substrates, with and without crc perturbation.
Hypothesis: vanAB expression is strongly substrate-responsive and further tuned by catabolite repression during mixed-substrate growth.
Type: transcriptional regulation analysis
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