Schizosaccharomyces pombe cps1 (SPAC24C9.08) encodes a vacuolar M20A metallopeptidase with conserved carboxypeptidase activity inferred from fungal ancestry. Its predicted zinc-dependent catalytic domain is associated with an N-terminal membrane anchor, and the protein enters the vacuolar sorting pathway. Cleavage of peptide substrates and amino-acid recycling in the vacuole are its inferred principal functions; its precise substrate range and native proteolytic maturation remain incompletely characterized. This protein is distinct from the glucan synthase Bgs1, which has also been called Cps1.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000324 fungal-type vacuole | HDA PMID:16823372 ORFeome cloning and global analysis of protein localization ... | ACCEPT | Summary: Localization to fungal-type vacuole is supported by high-throughput YFP tagging study. This is direct experimental evidence for localization. Supporting Evidence: PMID:16823372 we determined the localization of 4,431 proteins, corresponding to approximately 90% of the fission yeast proteome, by tagging each ORF with the yellow fluorescent protein |
| GO:0000328 fungal-type vacuole lumen | IBA GO_REF:0000033 | ACCEPT | Summary: Vacuolar-lumen localization is a credible inherited fungal CPS property. Reason: The exact target descends from the vacuolar CPS IBD. ScCPS1 membrane-bound precursors mature to a soluble lumenal enzyme (PMID:1569061). Target SpCPS vacuolar routing is independently observed using an engineered Ub-GFP fusion (PMID:17660439), while GFP alone perturbs sorting. That experiment does not prove native cleavage, but absence of a native processing assay does not invalidate the conserved localization inference; a membrane precursor and lumenal mature pool can coexist. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000865917 SUPPORTS TRANSFER Current PTHR45962 v19 exact O13968 leaf PTN000110806 descends from this fungal CPS node; the corresponding IBD is verified. No loss event is recovered on the target path. Mammalian PM20D1 activity is a separate functional comparison. Supporting Evidence: PMID:1569061 The mature forms of carboxypeptidase yscS appeared soluble in the vacuolar lumen, while the precursor proteins accumulated tightly associated with the vacuolar membrane. PMID:17660439 A CPS1 homologue, SPAC24C9.08 (SpCPS), was amplified by PCR and subcloned into pTN54 PMID:17660439 However, GFP–SpCPS was not effectively sorted into vacuoles even in wild-type. |
| GO:0005774 vacuolar membrane | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt annotation based on subcellular location vocabulary. The protein is predicted to be a type II membrane protein. For S. cerevisiae CPS1, the precursor is membrane-associated before processing releases the mature enzyme. Reason: Consistent with UniProt membrane topology annotation and YFP localization data |
| GO:0005576 extracellular region | IEA GO_REF:0000117 | UNDECIDED | Summary: An additional extracellular pool is unresolved. Reason: Vacuolar localization is established but does not prove exclusive residence. The ARBA source does not identify a direct SpCPS secretion experiment, and the mammalian PM20D1 secretion result cannot simply be assigned to this fungal protein. Retain uncertainty pending substrate/localization adjudication rather than infer absence solely from the main compartment. Supporting Evidence: PMID:17660439 A CPS1 homologue, SPAC24C9.08 (SpCPS), was amplified by PCR and subcloned into pTN54 PMID:17660439 However, GFP–SpCPS was not effectively sorted into vacuoles even in wild-type. |
| GO:0004180 carboxypeptidase activity | IBA GO_REF:0000033 | ACCEPT | Summary: Conserved vacuolar carboxypeptidase function supports carboxypeptidase activity. Reason: The fungal CPS ancestral node is present on the actual target lineage and is grounded in experimentally characterized ScCPS1. Target vacuolar localization and M20A architecture agree with that inference. Mammalian PM20D1 has additional/different substrate chemistry, but family diversity alone is not target-specific evidence of loss. Retain conserved peptide hydrolysis and vacuolar protein breakdown while leaving exact substrate preferences and additional nonpeptide activities open. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000865917 SUPPORTS TRANSFER Current PTHR45962 v19 exact O13968 leaf PTN000110806 descends from this fungal CPS node; the corresponding IBD is verified. No loss event is recovered on the target path. Mammalian PM20D1 activity is a separate functional comparison. Supporting Evidence: PMID:2026161 Chromosomal disruption of the CPS1 gene completely abolishes carboxypeptidase yscS activity. PMID:17660439 A CPS1 homologue, SPAC24C9.08 (SpCPS), was amplified by PCR and subcloned into pTN54 PMID:17660439 However, GFP–SpCPS was not effectively sorted into vacuoles even in wild-type. |
| GO:0004180 carboxypeptidase activity | IEA GO_REF:0000043 | ACCEPT | Summary: Conserved vacuolar carboxypeptidase function supports carboxypeptidase activity. Reason: The fungal CPS ancestral node is present on the actual target lineage and is grounded in experimentally characterized ScCPS1. Target vacuolar localization and M20A architecture agree with that inference. Mammalian PM20D1 has additional/different substrate chemistry, but family diversity alone is not target-specific evidence of loss. Retain conserved peptide hydrolysis and vacuolar protein breakdown while leaving exact substrate preferences and additional nonpeptide activities open. Supporting Evidence: PMID:2026161 Chromosomal disruption of the CPS1 gene completely abolishes carboxypeptidase yscS activity. PMID:17660439 A CPS1 homologue, SPAC24C9.08 (SpCPS), was amplified by PCR and subcloned into pTN54 PMID:17660439 However, GFP–SpCPS was not effectively sorted into vacuoles even in wild-type. |
| GO:0004181 metallocarboxypeptidase activity | IEA GO_REF:0000002 | ACCEPT | Summary: Conserved vacuolar carboxypeptidase function supports metallocarboxypeptidase activity. Reason: The fungal CPS ancestral node is present on the actual target lineage and is grounded in experimentally characterized ScCPS1. Target vacuolar localization and M20A architecture agree with that inference. Mammalian PM20D1 has additional/different substrate chemistry, but family diversity alone is not target-specific evidence of loss. Retain conserved peptide hydrolysis and vacuolar protein breakdown while leaving exact substrate preferences and additional nonpeptide activities open. Supporting Evidence: PMID:2026161 Chromosomal disruption of the CPS1 gene completely abolishes carboxypeptidase yscS activity. PMID:17660439 A CPS1 homologue, SPAC24C9.08 (SpCPS), was amplified by PCR and subcloned into pTN54 PMID:17660439 However, GFP–SpCPS was not effectively sorted into vacuoles even in wild-type. |
| GO:0051603 obsolete proteolysis involved in protein catabolic process | IBA GO_REF:0000033 | MODIFY | Summary: The inherited protein-catabolic function is retained using an active GO term. Reason: Live QuickGO confirms GO:0051603 was obsoleted as a precomposed term. Current PAINT already records protein catabolic process at the same fungal node. This is an ontology update, not rejection of conserved peptide hydrolysis; source term and evidence remain unchanged. Propagation Review Root cause: TERM SCOPING PROBLEM Sources checked: PANTHER:PTN000865917 SUPPORTS TRANSFER The current IBD at this exact ancestral node uses GO:0030163 protein catabolic process; frozen GO:0051603 is obsolete. Proposed replacements: protein catabolic process Supporting Evidence: PMID:2026161 Chromosomal disruption of the CPS1 gene completely abolishes carboxypeptidase yscS activity. |
| GO:0006508 proteolysis | IEA GO_REF:0000043 | ACCEPT | Summary: Conserved vacuolar carboxypeptidase function supports proteolysis. Reason: The fungal CPS ancestral node is present on the actual target lineage and is grounded in experimentally characterized ScCPS1. Target vacuolar localization and M20A architecture agree with that inference. Mammalian PM20D1 has additional/different substrate chemistry, but family diversity alone is not target-specific evidence of loss. Retain conserved peptide hydrolysis and vacuolar protein breakdown while leaving exact substrate preferences and additional nonpeptide activities open. Supporting Evidence: PMID:2026161 Chromosomal disruption of the CPS1 gene completely abolishes carboxypeptidase yscS activity. PMID:17660439 A CPS1 homologue, SPAC24C9.08 (SpCPS), was amplified by PCR and subcloned into pTN54 PMID:17660439 However, GFP–SpCPS was not effectively sorted into vacuoles even in wild-type. |
| GO:0007039 protein catabolic process in the vacuole | NAS GO_REF:0000051 | ACCEPT | Summary: Conserved vacuolar carboxypeptidase function supports protein catabolic process in the vacuole. Reason: The fungal CPS ancestral node is present on the actual target lineage and is grounded in experimentally characterized ScCPS1. Target vacuolar localization and M20A architecture agree with that inference. Mammalian PM20D1 has additional/different substrate chemistry, but family diversity alone is not target-specific evidence of loss. Retain conserved peptide hydrolysis and vacuolar protein breakdown while leaving exact substrate preferences and additional nonpeptide activities open. Supporting Evidence: PMID:2026161 Chromosomal disruption of the CPS1 gene completely abolishes carboxypeptidase yscS activity. PMID:17660439 A CPS1 homologue, SPAC24C9.08 (SpCPS), was amplified by PCR and subcloned into pTN54 PMID:17660439 However, GFP–SpCPS was not effectively sorted into vacuoles even in wild-type. |
| GO:0006520 amino acid metabolic process | IEA GO_REF:0000117 | ACCEPT | Summary: ARBA annotation. Whether carboxypeptidase or aminoacylase, the enzyme would be involved in amino acid metabolism. This general annotation is likely correct. Reason: Both carboxypeptidase and aminoacylase activities involve amino acid metabolism. This general term is appropriate regardless of specific enzymatic mechanism. |
| GO:0006629 lipid metabolic process | IEA GO_REF:0000117 | UNDECIDED | Summary: Additional N-acyl-amino-acid chemistry remains unresolved for SpCPS. Reason: Mammalian PM20D1 catalyzes reversible N-acyl-amino-acid formation/hydrolysis. This supplies a family-level hypothesis for lipid metabolism and nonpeptide C-N chemistry, not proof of either retention or loss in fungal CPS. Peptide and nonpeptide hydrolysis need not be mutually exclusive. GO:0043604 is also confirmed obsolete; an active product-specific biosynthetic replacement cannot be selected before the target reaction is established. Supporting Evidence: PMID:27374330 catalyzing both the condensation of fatty acids and amino acids to generate N-acyl amino acids and also the reverse hydrolytic reaction. |
| GO:0008233 peptidase activity | IEA GO_REF:0000043 | ACCEPT | Summary: Conserved vacuolar carboxypeptidase function supports peptidase activity. Reason: The fungal CPS ancestral node is present on the actual target lineage and is grounded in experimentally characterized ScCPS1. Target vacuolar localization and M20A architecture agree with that inference. Mammalian PM20D1 has additional/different substrate chemistry, but family diversity alone is not target-specific evidence of loss. Retain conserved peptide hydrolysis and vacuolar protein breakdown while leaving exact substrate preferences and additional nonpeptide activities open. Supporting Evidence: PMID:2026161 Chromosomal disruption of the CPS1 gene completely abolishes carboxypeptidase yscS activity. PMID:17660439 A CPS1 homologue, SPAC24C9.08 (SpCPS), was amplified by PCR and subcloned into pTN54 PMID:17660439 However, GFP–SpCPS was not effectively sorted into vacuoles even in wild-type. |
| GO:0016787 hydrolase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Both carboxypeptidase and aminoacylase activities are hydrolase activities. This general term is appropriate regardless of specific substrate. Reason: General parent term appropriate for M20 family enzymes |
| GO:0016810 hydrolase activity, acting on carbon-nitrogen (but not peptide) bonds | IEA GO_REF:0000117 | UNDECIDED | Summary: Additional N-acyl-amino-acid chemistry remains unresolved for SpCPS. Reason: Mammalian PM20D1 catalyzes reversible N-acyl-amino-acid formation/hydrolysis. This supplies a family-level hypothesis for lipid metabolism and nonpeptide C-N chemistry, not proof of either retention or loss in fungal CPS. Peptide and nonpeptide hydrolysis need not be mutually exclusive. GO:0043604 is also confirmed obsolete; an active product-specific biosynthetic replacement cannot be selected before the target reaction is established. Supporting Evidence: PMID:27374330 catalyzing both the condensation of fatty acids and amino acids to generate N-acyl amino acids and also the reverse hydrolytic reaction. |
| GO:0043604 obsolete amide biosynthetic process | IEA GO_REF:0000117 | UNDECIDED | Summary: Additional N-acyl-amino-acid chemistry remains unresolved for SpCPS. Reason: Mammalian PM20D1 catalyzes reversible N-acyl-amino-acid formation/hydrolysis. This supplies a family-level hypothesis for lipid metabolism and nonpeptide C-N chemistry, not proof of either retention or loss in fungal CPS. Peptide and nonpeptide hydrolysis need not be mutually exclusive. GO:0043604 is also confirmed obsolete; an active product-specific biosynthetic replacement cannot be selected before the target reaction is established. Supporting Evidence: PMID:27374330 catalyzing both the condensation of fatty acids and amino acids to generate N-acyl amino acids and also the reverse hydrolytic reaction. |
| GO:0043605 obsolete amide catabolic process | IEA GO_REF:0000117 | MODIFY | Summary: The obsolete broad amide-catabolism term can be expressed as proteolysis. Reason: Live QuickGO confirms GO:0043605 is obsolete. Peptide bonds are amide bonds, and the supported conserved peptide-hydrolysis role supplies the evidence-backed active replacement. This does not assume that the additional nonpeptide substrate hypothesis is false. Proposed replacements: proteolysis Supporting Evidence: PMID:2026161 Chromosomal disruption of the CPS1 gene completely abolishes carboxypeptidase yscS activity. |
| GO:0046872 metal ion binding | IEA GO_REF:0000043 | ACCEPT | Summary: M20 family enzymes bind zinc ions. This is well-supported by domain architecture. Reason: M20 family enzymes require metal ions for catalysis Supporting Evidence: file:SCHPO/cps1/cps1-uniprot.txt Note=Binds 2 Zn(2+) ions per subunit. file:SCHPO/cps1/cps1-deep-research-cyberian.md The M20 family employs a "co-catalytic" mechanism involving two zinc ions per monomer at the active site center |
| GO:1990845 adaptive thermogenesis | IEA GO_REF:0000117 | UNDECIDED | Summary: Adaptive thermogenesis is not established for SpCPS. Reason: The term means regulated heat production in response to environmental changes; it is not definitionally limited to brown adipocytes. Mammalian PM20D1-dependent uncoupling does not itself establish a corresponding regulated fungal pathway, while a lack of adipose tissue is not a sufficient refutation. The target pathway and contribution remain unresolved and are included in focused adjudication. Supporting Evidence: PMID:27374330 N-acyl amino acids directly bind mitochondria and function as endogenous uncouplers of UCP1-independent respiration. |
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Download this section (compressed HTML)Q: What peptide and nonpeptide N-acyl-amino-acid substrates does native SpCPS hydrolyze, and is measurable reverse amidation physiologically relevant?
Q: How is native SpCPS processed and partitioned between vacuolar membrane and lumen, and is any extracellular pool condition dependent?
Experiment: Express and purify recombinant SPAC24C9.08 and assay for carboxypeptidase activity using Cbz-Gly-Leu as substrate. Compare activity to S. cerevisiae CPS1. Test substrate specificity with various dipeptides to determine P1 preference.
Hypothesis: SPAC24C9.08 has carboxypeptidase activity similar to S. cerevisiae CPS1
Type: biochemical enzyme assay
Experiment: Assay purified SPAC24C9.08 for hydrolysis of N-acyl amino acids (e.g., N-acetyl- L-methionine, N-oleoyl-phenylalanine) and for synthase activity (condensation of fatty acids with amino acids).
Hypothesis: SPAC24C9.08 has aminoacylase/N-acyl amino acid hydrolase activity like PM20D1
Type: biochemical enzyme assay
Experiment: Generate SPAC24C9.08 deletion strain and assess vacuolar proteolysis using autophagy markers during nitrogen starvation. Compare to wild-type and to cpy1 (carboxypeptidase Y) deletion strains.
Hypothesis: SPAC24C9.08 deletion affects vacuolar proteolysis in S. pombe
Type: genetic analysis
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