hutH (PA5098) encodes histidine ammonia-lyase (histidase, EC 4.3.1.3), the first enzyme of the histidine utilization (hut) pathway of Pseudomonas aeruginosa. It is a cytoplasmic, homotetrameric member of the PAL/histidase (aromatic amino acid ammonia-lyase) family. Its catalytic electrophile is a 4-methylidene-imidazol-5-one (MIO) cofactor, formed autocatalytically from the internal Ala142-Ser143-Gly144 tripeptide. The enzyme eliminates ammonia from L-histidine to form trans-urocanate, which is both the substrate of urocanase (HutU) and the physiological inducer of the hut operons, acting through the HutC repressor. Histidase synthesis is induced by urocanate and strongly subject to catabolite and nitrogen control. The partially purified P. aeruginosa enzyme is about 210 kDa, with a Km for histidine near 2 mM and sigmoid kinetics under some conditions. The hut locus also encodes a second, divergent HutH-like protein (PA5093) that lacks the MIO-forming residues; PA5098 is the functional histidase.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003824 catalytic activity | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO from the L-aspartase-like fold superfamily (IPR008948). Reason: True but uninformative. The specific activity, histidine ammonia-lyase activity (GO:0004397), is an is_a descendant already carried by the gene. Proposed replacements: histidine ammonia-lyase activity Supporting Evidence: file:PSEAE/hutH/hutH-uniprot.txt Reaction=L-histidine = trans-urocanate + NH4(+) |
| GO:0004397 histidine ammonia-lyase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN000796665, the bacterial histidase node of PTHR10362. Its IBD is seeded only by this gene's PseudoCAP row. The target appearing in its own WITH/FROM is expected, not circular. Reason: PA5098 retains the MIO-forming Ala-Ser-Gly (142-144) and the PAL/histidase active-site signature, is 82% identical to the structurally characterised P. putida histidase, and the P. aeruginosa enzyme activity was measured directly. It is inside the clade that has this function. The node does over-propagate to PA5093, a PAO1 in-paralog that lacks the MIO motif; that is recorded as a source defect and does not affect this gene. Supporting Evidence: file:PSEAE/hutH/hutH-uniprot.txt Contains an active site 4-methylidene-imidazol-5-one (MIO), which is formed autocatalytically by cyclization and dehydration of residues Ala-Ser-Gly. PMID:4290562 The molecular weight of partially purified histidase was estimated at 210,000 by sucrose gradient centrifugation. file:PSEAE/hutH/hutH-bioinformatics/RESULTS.md PA5098 (Q9HU85) carries the MIO-forming Ala-Ser-Gly tripeptide at 142-144 |
| GO:0004397 histidine ammonia-lyase activity | IEA GO_REF:0000120 | ACCEPT | Summary: HAMAP/InterPro (HutH, IPR005921)/EC/Rhea-based histidase activity. Reason: Same correct claim as the IBA and IMP rows. Supporting Evidence: file:PSEAE/hutH/hutH-uniprot.txt Belongs to the PAL/histidase family. PMID:10220322 the reactive electrophile is a 4-methylidene-imidazole-5-one, which is formed autocatalytically by cyclization and dehydration of residues 142-144 with the sequence Ala-Ser-Gly |
| GO:0004397 histidine ammonia-lyase activity | IMP PMID:4290562 Formation and operation of the histidine-degrading pathway i... | ACCEPT | Summary: PseudoCAP row from Lessie and Neidhardt 1967. It is the sole seed of the PTN000796665 histidase IBD (21 leaf IBAs) and one of 9 seeds of the PTN000796617 ammonia-lyase IBD (269 leaf IBAs). The paper assays P. aeruginosa histidase, partially purifies it, and reports its size and Km. Reason: Sound seed. The paper measures histidase activity of the P. aeruginosa enzyme directly. The partially purified enzyme (about 210 kDa) fits a tetramer of the 53.8 kDa PA5098 subunit. PA5098 is the only PAL/histidase-family protein in PAO1 that has the MIO motif, so the measured activity can be assigned to it even though the 1967 paper predates genes. The evidence is a direct enzyme assay rather than a hutH mutant (the mutants described are urocanase-deficient and regulatory), so IDA would be the better code; this is flagged to PseudoCAP. It does not change the soundness of the IBDs. Supporting Evidence: PMID:4290562 Its K(m) for histidine was 2 x 10(-3)m in tris(hydroxymethyl)aminomethane chloride buffer. PMID:4290562 The molecular weight of partially purified histidase was estimated at 210,000 by sucrose gradient centrifugation. file:PSEAE/hutH/hutH-bioinformatics/RESULTS.md It is the functional histidase of PAO1. |
| GO:0005737 cytoplasm | IEA GO_REF:0000120 | ACCEPT | Summary: HAMAP/InterPro cytoplasm location. HutH is a soluble enzyme acting on histidine taken up by the HutTh transporter. Reason: Histidase is a soluble cytoplasmic enzyme; this is where the core activity occurs. Supporting Evidence: file:PSEAE/hutH/hutH-uniprot.txt SUBCELLULAR LOCATION: Cytoplasm |
| GO:0006548 L-histidine catabolic process | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN000796665 (bacterial histidase clade), seeded only by this gene's PseudoCAP row. Reason: Histidase catalyses step I of histidine degradation, so the enzyme does part of the work of the process. The term is shared by both FIGLU branches of P. aeruginosa and is the right level for this step. HutH-type genes are genetically required for growth on histidine in the close relative P. fluorescens SBW25. Supporting Evidence: PMID:16475788 The first three steps for the degradation of histidine include the action of HutH, HutU, and HutI to convert L-histidine to N-formimino-L-glutamate. PMID:17717196 The mutant strain PBR801 (Ξ hutH 1 /H 2 ) was unable to grow on histidine, but could grow on urocanate, which confirms the histidase (histidine ammonia-lyase) function of hutH 1 and/or hutH 2 . |
| GO:0006548 L-histidine catabolic process | IEA GO_REF:0000120 | ACCEPT | Summary: HAMAP/InterPro HutH-family mapping to L-histidine catabolic process. Reason: Same correct claim as the IBA and IMP rows. Supporting Evidence: file:PSEAE/hutH/hutH-uniprot.txt N-formimidoyl-L-glutamate from L-histidine: step 1/3. |
| GO:0006548 L-histidine catabolic process | IMP PMID:4290562 Formation and operation of the histidine-degrading pathway i... | ACCEPT | Summary: PseudoCAP row from Lessie and Neidhardt 1967; sole seed of the PTN000796665 L-histidine catabolic process IBD (21 leaf IBAs). The paper characterizes histidase as the inducible first enzyme of P. aeruginosa histidine degradation. Reason: Sound seed. The enzyme assayed is the step I catalyst of the pathway the paper studies, and that enzyme is PA5098 (see the MF row). The evidence is better described as IDA than IMP, but the process claim is correct. Supporting Evidence: PMID:4290562 Histidine ammonia lyase (histidase), urocanase, and the capacity to degrade formiminoglutamate, which are respectively involved in steps I, II, and IV in the catabolism of histidine PMID:4290562 The results imply that urocanate is the physiological inducer of the histidine-degrading enzymes in P. aeruginosa. |
| GO:0016841 ammonia-lyase activity | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO from the PAL/histidase active-site signature (IPR022313). Reason: Correct but general. The specific activity, histidine ammonia-lyase activity (GO:0004397), is an is_a descendant already carried by the gene. Proposed replacements: histidine ammonia-lyase activity Supporting Evidence: file:PSEAE/hutH/hutH-uniprot.txt Reaction=L-histidine = trans-urocanate + NH4(+) |
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Download this section (compressed HTML)Q: PAO1 PA5093 (Q9HU90) is a HutH-like in-paralog lacking the MIO Ala-Ser-Gly motif, yet it inherits histidase IBAs from PTN000796665. Should PAINT add NOT annotations at its leaf, and are other members of PTN000796665 (e.g. Fusobacterium FN0791, or the extra Shewanella copies) also MIO-less HutH1-type proteins?
Q: What is the function, if any, of the conserved MIO-less HutH1-type paralog (PA5093) found in the hut loci of most Pseudomonas species?
Q: Does the full text of PMID:4290562 describe a histidase-deficient mutant (supporting IMP), or only enzyme assays (supporting IDA)?
Experiment: Delete PA5093 and PA5098 singly and together in PAO1, test growth on histidine versus urocanate, and assay histidase in extracts and with purified proteins, to confirm that PA5098 alone is the histidase and to probe any histidine-binding or regulatory role of the MIO-less PA5093.
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