nicF (PP_3941, UniProt Q88FY5) encodes maleamate amidohydrolase (EC 3.5.1.107), a metal-independent cysteine amidase of the isochorismatase superfamily. NicF hydrolyzes maleamate (maleamic acid) to maleate (maleic acid) plus ammonia, the nitrogen-removing step in the maleamate route of aerobic nicotinate (nicotinic acid) degradation in Pseudomonas putida KT2440. It acts downstream of NicD and upstream of NicE (maleate isomerase), which converts maleate to fumarate for entry into central metabolism. NicF is a soluble, intracellular catabolic enzyme encoded within the chromosomal nic gene cluster.
Definition: Catalysis of the reaction maleamate + H2O = maleate + NH4(+).
Justification: NicF has a characterized EC 3.5.1.107 substrate-level 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:1901848 nicotinate catabolic process | IDA PMID:18678916 Deciphering the genetic determinants for aerobic nicotinic a... | ACCEPT | Summary: Nicotinate catabolic process is the correct pathway annotation for NicF. Reason: NicF is the maleamate amidase in the characterized KT2440 nicotinate degradation cluster, acting downstream of NicD and upstream of NicE. The falcon deep research reaffirms the explicit placement of nicF in the aerobic nicotinate (maleamate) pathway and its required role for complete catabolism of nicotinate. Supporting Evidence: file:PSEPK/nicF/nicF-uniprot.txt aerobic nicotinate degradation pathway file:PSEPK/nicF/nicF-goa.tsv GO:1901848 nicotinate catabolic process PMID:18678916 responsible for the aerobic NA degradation file:PSEPK/nicF/nicF-deep-research-falcon.md NicF acts in the **maleamate pathway** of aerobic nicotinic acid degradation, downstream of NicC/NicX/NicD and upstream of **NicE** (maleate isomerase), which converts maleate to fumarate. |
| 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: The broad linear-amide hydrolase term captures the current GO molecular-function support for NicF. Reason: NicF hydrolyzes maleamate to maleate and ammonium; GO lacks a substrate-specific maleamate amidohydrolase term, so this parent is currently appropriate. The falcon deep research adds quantitative biochemical support (crude-extract specific activity of 19.3 umol/min/mg with an NH4+-coupled assay) and mechanistic context (isochorismatase-superfamily, metal-independent cysteine amidase with a Cys-Asp-Lys triad), all consistent with a linear-amide C-N hydrolase. Supporting Evidence: file:PSEPK/nicF/nicF-uniprot.txt Maleamate amidase that transforms maleamate into maleate file:PSEPK/nicF/nicF-uniprot.txt Reaction=maleamate + H2O = maleate + NH4(+) file:PSEPK/nicF/nicF-goa.tsv GO:0016811 hydrolase activity file:PSEPK/nicF/nicF-deep-research-falcon.md NicF is the maleamate (maleamic acid) amidohydrolase** of the *P. putida* KT2440 nicotinate degradation pathway, catalyzing hydrolysis of **maleamate/maleamic acid** to **maleate/maleic acid plus ammonia (NH3/NH4+)** file:PSEPK/nicF/nicF-deep-research-falcon.md NicF in KT2440 is predicted and supported to belong to the **isochorismatase superfamily**, which includes cysteine-dependent amide/ester hydrolases often featuring a **Cys–Asp–Lys** catalytic triad. |
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Download this section (compressed HTML)Q: Should GO add a substrate-specific maleamate amidohydrolase molecular-function term for NicF-family enzymes?
Experiment: Measure NicF kinetics on maleamate and related amides, and test rescue of nicF mutant growth on nicotinate.
Type: enzyme kinetics and complementation assay
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