Ppx (PA5241) is the exopolyphosphatase of Pseudomonas aeruginosa, a 506-residue member of the GppA/Ppx family (ASKHA fold) 41% identical to E. coli Ppx. It processively hydrolyses inorganic polyphosphate (polyP) from the chain ends, releasing orthophosphate, and is activated by K+, NH4+ and Zn2+. The same N-terminal active site also transfers phosphate from polyP to ADP to make ATP, at about a tenth of the hydrolase rate. Together with polyphosphate kinase (Ppk), encoded by the adjacent divergent gene, Ppx sets the cellular polyP pool. ppx is transcribed from a sigma54/NtrC-dependent promoter under nitrogen limitation and from a sigma70/PhoB-dependent promoter under phosphate limitation. Loss of Ppx disturbs polyP homeostasis and has pleiotropic effects on motility, pyocyanin, acyl-homoserine lactone and rhamnolipid production and biofilm formation. By similarity to E. coli Ppx it is a peripheral membrane protein.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004309 exopolyphosphatase activity | EXP PMID:26576296 Pseudomonas aeruginosa Exopolyphosphatase Is Also a Polyphos... | ACCEPT | Summary: UniProt EXP. Purified recombinant paPpx hydrolyses polyP (Km 1.3-11 uM, kcat 40-57 s-1 by chain length), activated by K+, NH4+ and Zn2+. Reason: Direct biochemical evidence for the core activity. Supporting Evidence: PMID:26576296 Pseudomonas aeruginosa exopolyphosphatase (paPpx; EC 3.6.1.11) catalyzes the hydrolysis of polyphosphates (polyP), producing polyPn-1 plus inorganic phosphate (Pi). |
| GO:0004309 exopolyphosphatase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from the PTN000768205 IBD (seeds ppx itself, the expected self-seed, and E. coli Ppx P0AFL6). The exopolyphosphatase term suits the Ppx subclade and the IBD stands on E. coli Ppx alone. Reason: Core activity, experimentally established for this protein and inherited within the Ppx clade. Supporting Evidence: PMID:26576296 Pseudomonas aeruginosa exopolyphosphatase (paPpx; EC 3.6.1.11) catalyzes the hydrolysis of polyphosphates (polyP), producing polyPn-1 plus inorganic phosphate (Pi). |
| GO:0004309 exopolyphosphatase activity | IDA PMID:10382967 The gene for an exopolyphosphatase of Pseudomonas aeruginosa... | ACCEPT | Summary: PseudoCAP IDA and a seed of PTN000768198 (pyrophosphatase activity) and PTN000768205 (exopolyphosphatase activity). The overproduced enzyme releases Pi from polyP of chain length 79 and 750, matching the polyP lost. Reason: Seed verdict SOUND; confirmed independently with purified enzyme and kinetics (PMID:26576296). Supporting Evidence: PMID:10382967 Thus ppx encodes an enzyme that has exopoly(P)ase activity. PMID:26576296 Pseudomonas aeruginosa exopolyphosphatase (paPpx; EC 3.6.1.11) catalyzes the hydrolysis of polyphosphates (polyP), producing polyPn-1 plus inorganic phosphate (Pi). |
| GO:0004309 exopolyphosphatase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined IEA (InterPro IPR022371, RHEA:21528, EC 3.6.1.11). Reason: Same claim as the experimental rows. Supporting Evidence: PMID:26576296 Pseudomonas aeruginosa exopolyphosphatase (paPpx; EC 3.6.1.11) catalyzes the hydrolysis of polyphosphates (polyP), producing polyPn-1 plus inorganic phosphate (Pi). |
| GO:0005886 plasma membrane | IEA GO_REF:0000044 | KEEP AS NON CORE | Summary: SubCell mapping from the UniProt "Cell membrane; peripheral" note, itself by similarity to E. coli Ppx. Reason: Plausible peripheral membrane location transferred from E. coli Ppx; not shown for this protein and peripheral to the activity. Supporting Evidence: file:PSEAE/ppx/ppx-uniprot.txt SUBCELLULAR LOCATION: Cell membrane {ECO:0000250|UniProtKB:P0AFL6}; Peripheral membrane protein {ECO:0000250|UniProtKB:P0AFL6}. |
| GO:0005886 plasma membrane | ISS GO_REF:0000024 | KEEP AS NON CORE | Summary: UniProt ISS from E. coli Ppx (P0AFL6), a peripheral inner-membrane protein. Reason: Reasonable transfer from a 41%-identical ortholog; no direct data; non-core. Supporting Evidence: file:PSEAE/ppx/ppx-uniprot.txt SUBCELLULAR LOCATION: Cell membrane {ECO:0000250|UniProtKB:P0AFL6}; Peripheral membrane protein {ECO:0000250|UniProtKB:P0AFL6}. |
| GO:0006793 phosphorus metabolic process | IEA GO_REF:0000002 | TOO GENERAL | Summary: Generic InterPro2GO process term. Reason: Entailed by the accepted GO:0006798 polyphosphate catabolic process. |
| GO:0006798 polyphosphate catabolic process | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN000768205, seeded by E. coli Ppx. Ppx hydrolyses polyP, which is polyP catabolism. Reason: Direct participation: the enzyme catalyses polyP breakdown. Supporting Evidence: PMID:26576296 Pseudomonas aeruginosa exopolyphosphatase (paPpx; EC 3.6.1.11) catalyzes the hydrolysis of polyphosphates (polyP), producing polyPn-1 plus inorganic phosphate (Pi). |
| GO:0006995 cellular response to nitrogen starvation | IDA PMID:24275100 Exopolyphosphatase of Pseudomonas aeruginosa is essential fo... | MARK AS OVER ANNOTATED | Summary: PseudoCAP IDA from PMID:24275100 (abstract only). The data behind it are promoter mapping and regulation: ppx is transcribed from an NtrC-activated sigma54 promoter under nitrogen limitation. Reason: Transcriptional induction alone does not show that Ppx takes part in the starvation response, and the same paper reports that Ppx levels stay constant across conditions. Over-annotation; full text not read, so no defect filed. Supporting Evidence: PMID:24275100 Under nitrogen-limiting conditions, its expression was controlled from a Ο(54)-dependent promoter activated by the response regulator NtrC. PMID:24275100 Molecular and physiological approaches used in this study indicated that P. aeruginosa maintains consistently proper levels of Ppx regardless of environmental conditions. |
| GO:0008976 polyphosphate kinase activity | EXP PMID:26576296 Pseudomonas aeruginosa Exopolyphosphatase Is Also a Polyphos... | KEEP AS NON CORE | Summary: UniProt EXP. Purified paPpx makes ATP from polyP and ADP at the same active site as the hydrolase, with kcat about 4 s-1, about a tenth of the hydrolase rate. Reason: Genuine secondary activity of the same active site (GO:0008976 is defined in the ATP-to-polyP direction but RHEA:19573 is reversible); the main activity is hydrolysis. Supporting Evidence: PMID:26576296 We show that paPpx is also a polyphosphate:ADP phosphotransferase and that the active site is the same as that one involved in the hydrolase activity. |
| GO:0008976 polyphosphate kinase activity | IEA GO_REF:0000120 | KEEP AS NON CORE | Summary: Rhea/EC IEA for polyphosphate kinase activity (EC 2.7.4.1). Reason: Same claim as the EXP row; secondary activity. Supporting Evidence: PMID:26576296 We show that paPpx is also a polyphosphate:ADP phosphotransferase and that the active site is the same as that one involved in the hydrolase activity. |
| GO:0009247 glycolipid biosynthetic process | IMP PMID:24275100 Exopolyphosphatase of Pseudomonas aeruginosa is essential fo... | REMOVE | Summary: PseudoCAP IMP from PMID:24275100: the ppx null mutant is altered in rhamnolipid production. Reason: Indirect phenotype. Ppx hydrolyses polyP and does not catalyse any step of rhamnolipid (glycolipid) synthesis, which is done by RhlA, RhlB and RhlC; the change follows from disturbed polyP/Pi homeostasis. Supporting Evidence: PMID:24275100 Results obtained from the ppx null mutant demonstrated that Ppx is involved in the production of virulence factors associated with both acute infection (e.g. motility-promoting factors, blue/green pigment production, C6-C12 quorum-sensing homoserine lactones) and chronic infection (e.g. rhamnolipids, biofilm formation). |
| GO:0009372 quorum sensing | IMP PMID:24275100 Exopolyphosphatase of Pseudomonas aeruginosa is essential fo... | REMOVE | Summary: PseudoCAP IMP from PMID:24275100: the ppx null mutant has altered C6-C12 homoserine lactone levels. Reason: Indirect phenotype. Ppx neither synthesises nor senses AHL signals; altered AHL levels in the mutant reflect pleiotropic effects of polyP/Pi imbalance. Supporting Evidence: PMID:24275100 Results obtained from the ppx null mutant demonstrated that Ppx is involved in the production of virulence factors associated with both acute infection (e.g. motility-promoting factors, blue/green pigment production, C6-C12 quorum-sensing homoserine lactones) and chronic infection (e.g. rhamnolipids, biofilm formation). |
| GO:0016036 cellular response to phosphate starvation | IDA PMID:24275100 Exopolyphosphatase of Pseudomonas aeruginosa is essential fo... | MARK AS OVER ANNOTATED | Summary: PseudoCAP IDA from PMID:24275100 (abstract only), based on PhoB-activated sigma70 transcription of ppx under Pi limitation. Reason: Expression control alone does not show Ppx participates in the phosphate starvation response; Ppx levels were reported as constant. Over-annotation; full text not read, so no defect filed. Supporting Evidence: PMID:24275100 However, under Pi limitation, the expression was controlled from a Ο(70) promoter, activated by PhoB. PMID:24275100 Molecular and physiological approaches used in this study indicated that P. aeruginosa maintains consistently proper levels of Ppx regardless of environmental conditions. |
| GO:0044010 single-species biofilm formation | IMP PMID:24275100 Exopolyphosphatase of Pseudomonas aeruginosa is essential fo... | KEEP AS NON CORE | Summary: PseudoCAP IMP: the ppx null mutant forms altered biofilm. Reason: Organism-level process term from a mutant phenotype, as GO conventionally annotates; pleiotropic and non-core. Supporting Evidence: PMID:24275100 Results obtained from the ppx null mutant demonstrated that Ppx is involved in the production of virulence factors associated with both acute infection (e.g. motility-promoting factors, blue/green pigment production, C6-C12 quorum-sensing homoserine lactones) and chronic infection (e.g. rhamnolipids, biofilm formation). |
| GO:0071977 bacterial-type flagellum-dependent swimming motility | IMP PMID:24275100 Exopolyphosphatase of Pseudomonas aeruginosa is essential fo... | KEEP AS NON CORE | Summary: PseudoCAP IMP: the ppx null mutant is altered in motility. Reason: Mutant-phenotype process annotation; pleiotropic and non-core. Supporting Evidence: PMID:24275100 Results obtained from the ppx null mutant demonstrated that Ppx is involved in the production of virulence factors associated with both acute infection (e.g. motility-promoting factors, blue/green pigment production, C6-C12 quorum-sensing homoserine lactones) and chronic infection (e.g. rhamnolipids, biofilm formation). |
| GO:0071978 bacterial-type flagellum-dependent swarming motility | IDA PMID:24275100 Exopolyphosphatase of Pseudomonas aeruginosa is essential fo... | KEEP AS NON CORE | Summary: PseudoCAP row coded IDA, but the abstract describes motility effects only from the ppx null mutant, so the evidence is probably mutant phenotype (IMP). Reason: Same standing as the swimming row; pleiotropic and non-core. The evidence code looks wrong but the full text was not read, so no defect is filed. Supporting Evidence: PMID:24275100 Results obtained from the ppx null mutant demonstrated that Ppx is involved in the production of virulence factors associated with both acute infection (e.g. motility-promoting factors, blue/green pigment production, C6-C12 quorum-sensing homoserine lactones) and chronic infection (e.g. rhamnolipids, biofilm formation). |
Loading supporting contentβ¦
Download this section (compressed HTML)Q: Is the polyP:ADP phosphotransferase activity of Ppx physiologically significant for ATP regeneration in P. aeruginosa, or a side reaction of the hydrolase site?
Q: Which of the ppx-mutant virulence phenotypes are due to polyP accumulation and which to altered Pi or ppGpp signalling?
Experiment: Compare ppx, ppk and ppx ppk mutants for polyP levels and the motility, AHL and rhamnolipid phenotypes, and complement ppx with a catalytically dead allele, to separate direct from indirect effects.
Loading supporting contentβ¦
Download this section (compressed HTML)Loading supporting contentβ¦
Download this section (compressed HTML)