nicD encodes N-formylmaleamate deformylase, an alpha/beta-hydrolase-fold enzyme in the aerobic nicotinate degradation cluster of Pseudomonas putida KT2440. NicD hydrolyzes N-formylmaleamate to maleamate and formate downstream of NicX, completing a key step in nicotinate catabolism.
Definition: Catalysis of the reaction N-formylmaleamate + H2O = maleamate + formate + H(+).
Justification: NicD has a characterized substrate-level deformylase reaction in nicotinate catabolism, while the current GO annotation can only use the broader linear-amide hydrolase parent.
Parent term: hydrolase activity, acting on carbon-nitrogen (but not peptide) bonds, in linear amides
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0016020 membrane | IEA GO_REF:0000118 | REMOVE | Summary: The generic membrane annotation is unsupported for NicD. Reason: The curated UniProt entry describes a soluble enzymatic role in nicotinate degradation and has no subcellular-location or transmembrane evidence supporting membrane localization. Falcon deep research confirms NicD is best annotated as cytosolic, assayed in soluble crude extracts, with no membrane segment, signal peptide, or export evidence reported. Supporting Evidence: file:PSEPK/nicD/nicD-uniprot.txt Deformylase that catalyzes the conversion of N-formylmaleamic file:PSEPK/nicD/nicD-deep-research-falcon.md Best current annotation is **cytosolic**: NicD was assayed in **soluble crude extracts** after heterologous expression; no membrane segment, signal peptide, or export evidence is reported in the cited sources. |
| GO:1901848 nicotinate catabolic process | IDA PMID:18678916 Deciphering the genetic determinants for aerobic nicotinic a... | ACCEPT | Summary: Nicotinate catabolic process is the correct pathway for NicD. Reason: The KT2440 nic cluster paper and UniProt pathway section place NicD in aerobic nicotinate degradation. Falcon deep research confirms NicD acts downstream of NicX and upstream of NicF in the maleamate pathway, and that disruption of nicD prevents growth on nicotinic acid as sole carbon source, establishing an essential pathway role. Supporting Evidence: file:PSEPK/nicD/nicD-uniprot.txt Deformylase that catalyzes the conversion of N-formylmaleamic PMID:18678916 Further conversion of N-formylmaleamic acid to formic and maleamic acid is catalyzed by the NicD protein file:PSEPK/nicD/nicD-uniprot.txt PATHWAY: Cofactor degradation; nicotinate degradation file:PSEPK/nicD/nicD-deep-research-falcon.md Disruption of **nicD** prevented growth on **nicotinic acid** as sole carbon source, supporting an essential pathway role in KT2440; the nic cluster as a cassette restored NA utilization in nondegrading backgrounds. |
| GO:0016811 hydrolase activity, acting on carbon-nitrogen (but not peptide) bonds, in linear amides | IDA PMID:18678916 Deciphering the genetic determinants for aerobic nicotinic a... | ACCEPT | Summary: Hydrolase activity acting on C-N bonds in linear amides captures the current GO molecular-function support for NicD deformylase chemistry. Reason: The exact deformylase activity lacks a specific GO term in the current annotations, but NicD hydrolyzes N-formylmaleamate to maleamate and formate through an amide-bond hydrolase reaction. Falcon deep research corroborates the deformylase reaction and the alpha/beta-hydrolase catalytic triad (S101-D125-H245) underpinning this molecular function. Supporting Evidence: file:PSEPK/nicD/nicD-uniprot.txt Deformylase that catalyzes the conversion of N-formylmaleamic PMID:18678916 Further conversion of N-formylmaleamic acid to formic and maleamic acid is catalyzed by the NicD protein file:PSEPK/nicD/nicD-deep-research-falcon.md NicD catalyzes **N-formylmaleamic acid (NFM) → maleamic acid + formic acid**; this corresponds to **EC 3.5.1.106** (N-formylmaleamate deformylase) and removes the N-formyl group generated after pyridine-ring cleavage. file:PSEPK/nicD/nicD-deep-research-falcon.md Site-directed mutagenesis supports **S101–D125–H245** as the catalytic triad; **S101A, D125A, H245A** abolished activity, whereas **E221A** retained ~**70%** of wild-type activity, arguing E221 is not the catalytic acid. file:PSEPK/nicD/nicD-uniprot.txt Reaction=N-formylmaleamate + H2O = maleamate + formate |
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Download this section (compressed HTML)Q: Should GO add a substrate-specific N-formylmaleamate deformylase molecular-function term for NicD-family enzymes?
Suggested experts: GO molecular function editors, Nicotinate degradation experts
Experiment: Compare wild-type and catalytic-triad mutant NicD enzymes for N-formylmaleamate hydrolysis and rescue of nicD mutant growth on nicotinate.
Type: enzyme and complementation assay
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