PhzC is the class-II (type II) 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase encoded in each of the two phenazine biosynthetic operons of Pseudomonas aeruginosa PAO1 (phzC1 at PA4212 and the identical phzC2 at PA1901). It condenses phosphoenolpyruvate with D-erythrose 4-phosphate, the committed first step of the shikimate pathway, and so feeds carbon towards chorismate, the substrate from which PhzE begins phenazine assembly. Unlike the housekeeping DAH7PS of the same organism (PA2843), the phz-operon enzyme is a structurally distinct, allosterically unregulated homotetramer: it is not inhibited by tryptophan, tyrosine, phenylalanine, phenazine-1-carboxylate or pyocyanin, which lets phenazine production continue when primary aromatic amino acid biosynthesis is switched off. In the related phenazine producer Pseudomonas chlororaphis, deleting phzC nearly abolishes phenazine-1-carboxamide output while deleting the housekeeping aro genes barely affects it, confirming that this duplicated DAH7PS is dedicated to secondary metabolism.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0002047 phenazine biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN000481936, a Proteobacteria node of PTHR21337 with five leaves, sole-seeded by this gene's own PseudoCAP IDA row. The claim is right for phzC1, but the node is a generic DAHP synthase clade: its other members are Helicobacter pylori HP_0134, Xanthomonas dhs1, Bradyrhizobium blr3778 and P. aeruginosa's own housekeeping DAH7PS PA2843, none of which belongs to a phenazine pathway. Reason: Correct for this gene, which is the phenazine operon's own DAH7PS and supplies the pathway's shikimate flux. Kept rather than removed because the claim is true of the target; the problem is the node it is asserted at, recorded as a GO_CENTRAL defect so the other four leaves stop inheriting it. Propagation Review Root cause: PROPAGATION BAD Failure modes: WRONG ORTHOLOG OR PARALOG LINEAGE OR TAXON MISMATCH Sources checked: PANTHER:PTN000481936 Β· PTN000481936 SOURCE BAD Wrong level for a pathway-specific term. The node groups the phz-cluster DAH7PS with full-length type II DAH7PSs, including the target organism's own paralogue PA2843 (Q9I000, 448 aa), which PMID:30242059 places outside the phzA-G cluster and in primary metabolism on the basis of its structure, its retained alpha2a/alpha2b helices and its inhibition by aromatic amino acids. UniProtKB:Q7DC82 Β· phzC1 (PA4212 / PA1901) SUPPORTS TRANSFER Sole seed, and the gene appearing in its own with/from is expected: the IDA row is the descendant evidence the PAINT curator used. The seed claim is independently supported by the enzyme's kinetic and allosteric characterization and by phzC deletion in P. chlororaphis. Supporting Evidence: PMID:30242059 Consistent with this is the observation made during functional characterisation that PaeDAH7PSPA1901 is insensitive to allosteric regulation by aromatic amino acids, confirming that PaeDAH7PSPA1901 functions primarily within secondary metabolism. file:PSEAE/phzC1/phzC1-bioinformatics/RESULTS.md A QuickGO check of the paralogue PA2843 (Q9I000) shows it carrying `involved_in GO:0002047` by IBA |
| GO:0002047 phenazine biosynthetic process | IDA PMID:11591691 Functional analysis of genes for biosynthesis of pyocyanin a... | ACCEPT | Summary: PseudoCAP IDA, the sole seed of the PTN000481936 IBD. Mavrodi et al. cloned the two seven-gene operons and showed that each is sufficient for phenazine-1-carboxylic acid production in phenazine-non-producing pseudomonads; the same IDA row was applied to the other genes of the operons. The evidence is therefore operon-level rather than gene-by-gene, which is worth noting beside an IDA code, but PhzC's own contribution is independently established. Reason: Core process. PhzC is the operon's own DAH7PS and performs the first shikimate-pathway step that supplies chorismate for the pathway; it is kinetically competent, is not feedback-inhibited by aromatic amino acids or phenazines, and its deletion in P. chlororaphis collapses phenazine output while housekeeping aro deletions do not. Supporting Evidence: PMID:11591691 Functional studies of phenazine-nonproducing strains of fluorescent pseudomonads indicated that each of the biosynthetic operons from P. aeruginosa is sufficient for production of a single compound, phenazine-1-carboxylic acid (PCA). PMID:35053084 After the complementation of gene phzC in the corresponding DAHP synthase-deficient strain, the production of PCN was significantly restored. PMID:30242059 Apparent KM values for PaeDAH7PSPA1901 for PEP and E4P were determined to be 17 Β± 1 and 16 Β± 3 ΞΌM respectively (Table 1). |
| GO:0003849 3-deoxy-7-phosphoheptulonate synthase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from the PTHR21337 family root PTN000481914 (186 leaves), with two experimental seeds: Arabidopsis AT4G39980 and Mycobacterium tuberculosis DAH7PS (O53512). This is the family's defining activity. Reason: Core molecular function, confirmed directly for this protein by steady-state kinetics on the purified enzyme, so the phylogenetic transfer is not the only support. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000481914 Β· PTN000481914 (PTHR21337 root) SUPPORTS TRANSFER Family-root IBD for the DAHP synthase reaction, appropriate for every member of the family. UniProtKB:O53512 Β· M. tuberculosis DAH7PS SUPPORTS TRANSFER Characterized type II DAH7PS, the structural comparator used in the P. aeruginosa structural study. Supporting Evidence: PMID:30242059 Apparent KM values for PaeDAH7PSPA1901 for PEP and E4P were determined to be 17 Β± 1 and 16 Β± 3 ΞΌM respectively (Table 1). file:PSEAE/phzC1/phzC1-uniprot.txt DR Pfam; PF01474; DAHP_synth_2; 1. |
| GO:0003849 3-deoxy-7-phosphoheptulonate synthase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Mapping from the InterPro class-II DAHP synthase signature and the EC number on the UniProt entry; agrees with the IBA row and with the measured kinetics. Reason: Correct and as specific as the reaction allows. Supporting Evidence: file:PSEAE/phzC1/phzC1-uniprot.txt Reaction=D-erythrose 4-phosphate + phosphoenolpyruvate + H2O = 7- |
| GO:0009073 aromatic amino acid family biosynthetic process | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: InterPro-to-GO mapping that attaches the generic shikimate-pathway role to every class-II DAHP synthase. For this paralogue the shikimate flux it generates is used by the phenazine pathway, not by aromatic amino acid biosynthesis, and the enzyme lacks the allosteric control that the primary-metabolism enzymes have. Reason: A domain-level inference that does not hold for this gene copy. The organism's aromatic amino acid biosynthesis is served by the housekeeping DAH7PS PA2843 and the aro genes; the phz-operon enzyme is insensitive to feedback by Trp, Tyr and Phe, which is the hallmark of its secondary-metabolism role. The chemistry is shared, so the activity term GO:0003849 is kept, but the primary-metabolism process is not this protein's. Supporting Evidence: PMID:30242059 At concentrations up to 200 ΞΌM Trp, Tyr, Phe, phenazine or PCA, PaeDAH7PSPA1901 activity was found to be comparable with that observed in the absence of aromatic amino acids or secondary metabolites PMID:35053084 As shown in Figure 4, no morphological difference was found, whereas phzC mutant strain HT66ΞphzC displayed a reduction in PCN production. |
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Download this section (compressed HTML)Q: Should the GO:0002047 IBD be moved from PTN000481936 to a node containing only phz-cluster DAH7PS proteins, so that Helicobacter pylori HP_0134, Xanthomonas dhs1, Bradyrhizobium blr3778 and P. aeruginosa's housekeeping PA2843 stop inheriting phenazine biosynthetic process?
Q: Do the two identical phzC copies (PA1901 and PA4212) contribute equally, and is either of them dispensable when the other operon is expressed? The GOA entry merges them, so the question cannot currently be asked of the annotations.
Experiment: Make single and double phzC deletions in PAO1 (PA1901, PA4212 and both) and measure PCA, pyocyanin and intracellular chorismate, with complementation by the housekeeping PA2843 enzyme as a control for whether the secondary-metabolism-specific, feedback-insensitive DAH7PS is required or merely sufficient. This would convert the current operon-level IDA into gene-level evidence in P. aeruginosa itself.
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