PA1242 encodes SprP, one of the two subtilisin-like (peptidase S8) serine proteases of Pseudomonas aeruginosa PAO1. The 590-residue precursor has an N-terminal signal peptide, a Pseudomonas-specific domain of unknown function and a C-terminal peptidase S8 domain with an unusual Glu262/His299/Ser499 catalytic triad. Recombinant SprP is autocatalytically processed to an active form of about 26 kDa that hydrolyses casein and the subtilisin substrate Suc-AAPF-pNA, is inhibited by AEBSF and TPCK and does not need calcium; the S499A variant is inactive. Deletion of sprP has pleiotropic effects, with altered expression of about 200 genes, abolished swimming, swarming and twitching motility, increased cell aggregation, biofilm formation and pyoverdine production, and reduced anaerobic growth. sprP transcription rises at 43 C, suggesting a role in the heat shock response. Its natural substrates are unknown.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004252 serine-type endopeptidase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN002459139, seeded by B. subtilis AprE (P04189), Plasmodium SUB1 (Q8I430) and this gene. Reason: Serine-type endopeptidase activity is shown for purified SprP and the IBD stands on the other seeds. Supporting Evidence: PMID:26175208 Our results confirm the classification of SprP as a serine protease PMID:26175208 The activity of SprP was strongly reduced by AEBSF and TPCK. PMID:24376018 SprP variant S499A which had the catalytic serine replaced by alanine also did not show protease activity |
| GO:0004252 serine-type endopeptidase activity | IDA PMID:26175208 Functional expression, purification, and biochemical propert... | ACCEPT | Summary: PseudoCAP IDA seed of PTN002459139 from the purified-enzyme paper. Reason: Purified SprP hydrolyses casein and is inhibited by the serine protease inhibitors AEBSF and TPCK; seed verdict SOUND. Supporting Evidence: PMID:26175208 Our results confirm the classification of SprP as a serine protease PMID:26175208 The activity of SprP was strongly reduced by AEBSF and TPCK. |
| GO:0004252 serine-type endopeptidase activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro subtilase superfamily mapping. Reason: Agrees with the experimental data. Supporting Evidence: file:PSEAE/PA1242/PA1242-uniprot.txt PF00082; Peptidase_S8 PMID:26175208 Our results confirm the classification of SprP as a serine protease |
| GO:0006508 proteolysis | IBA GO_REF:0000033 | ACCEPT | Summary: IBA proteolysis from PTN002459139. Reason: SprP is an active protease that also processes itself. Supporting Evidence: PMID:26175208 SprP is autocatalytically activated by proteolysis |
| GO:0006508 proteolysis | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro mapping to proteolysis. Reason: As for the IBA row. Supporting Evidence: PMID:26175208 SprP is autocatalytically activated by proteolysis |
| GO:0008233 peptidase activity | IDA PMID:24376018 Subtilase SprP exerts pleiotropic effects in Pseudomonas aer... | TOO GENERAL | Summary: IDA peptidase activity from the first SprP paper (skim milk, Suc-AAPF-pNA). Reason: Entailed by GO:0004252 serine-type endopeptidase activity, accepted here. Supporting Evidence: PMID:24376018 SprP variant S499A which had the catalytic serine replaced by alanine also did not show protease activity |
| GO:0008236 serine-type peptidase activity | IEA GO_REF:0000002 | TOO GENERAL | Summary: InterPro mapping to serine-type peptidase activity. Reason: Entailed by GO:0004252 serine-type endopeptidase activity, accepted here. |
| GO:0043107 type IV pilus-dependent motility | IMP PMID:24376018 Subtilase SprP exerts pleiotropic effects in Pseudomonas aer... | MARK AS OVER ANNOTATED | Summary: IMP from the deletion mutant, which lost twitching motility. Reason: The deletion abolishes swimming, swarming and twitching alike and changes expression of 218 genes. SprP is a protease of unknown substrate, not a part of the pilus machinery; the phenotype is indirect and does not show that SprP takes part in pilus-dependent motility. Supporting Evidence: PMID:24376018 Examination of P. aeruginosa ΞsprP for its swimming, swarming, and twitching motility revealed that all three types of motility were virtually abolished |
| GO:0071977 bacterial-type flagellum-dependent swimming motility | IMP PMID:24376018 Subtilase SprP exerts pleiotropic effects in Pseudomonas aer... | MARK AS OVER ANNOTATED | Summary: IMP from the deletion mutant, which lost swimming motility. Reason: As for twitching; a pleiotropic, indirect mutant phenotype. Supporting Evidence: PMID:24376018 Examination of P. aeruginosa ΞsprP for its swimming, swarming, and twitching motility revealed that all three types of motility were virtually abolished |
| GO:0071978 bacterial-type flagellum-dependent swarming motility | IMP PMID:24376018 Subtilase SprP exerts pleiotropic effects in Pseudomonas aer... | MARK AS OVER ANNOTATED | Summary: IMP from the deletion mutant, which lost swarming motility. Reason: As for twitching; a pleiotropic, indirect mutant phenotype. Supporting Evidence: PMID:24376018 Examination of P. aeruginosa ΞsprP for its swimming, swarming, and twitching motility revealed that all three types of motility were virtually abolished |
| GO:1900232 negative regulation of single-species biofilm formation on inanimate substrate | IMP PMID:24376018 Subtilase SprP exerts pleiotropic effects in Pseudomonas aer... | KEEP AS NON CORE | Summary: IMP from a sixfold increase in biofilm in the deletion mutant, reversed by complementation. Reason: A complemented mutant phenotype supports a regulatory role of SprP in biofilm formation; the mechanism is unknown, so it is kept as non-core. Supporting Evidence: PMID:24376018 P. aeruginosa ΞsprP showed an almost sixfold increased biofilm formation as compared to the wild-type strain P. aeruginosa PAO1 |
| GO:1900377 negative regulation of secondary metabolite biosynthetic process | IMP PMID:24376018 Subtilase SprP exerts pleiotropic effects in Pseudomonas aer... | MODIFY | Summary: IMP from fivefold higher pyoverdine in the deletion mutant, reversed by complementation. Reason: The secondary metabolite measured is the siderophore pyoverdine, so the specific term is negative regulation of siderophore biosynthetic process. Non-core; mechanism unknown. Proposed replacements: negative regulation of siderophore biosynthetic process Supporting Evidence: PMID:24376018 The culture supernatant of P. aeruginosa ΞsprP contained almost fivefold the amount of pyoverdine as compared to the wild-type supernatant |
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Experiment: Compare the proteomes and secretomes of wild type, delta-sprP and an S499A knock-in strain to find SprP substrates and to separate catalytic from structural effects on motility and biofilm.
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