CWC27 is the human ortholog of yeast Cwc27 and is a spliceosome-associated protein. As part of the spliceosome it functions in pre-mRNA splicing. It is recruited during activation as a component of the activated (Bact) spliceosome and is one of the first proteins released as the spliceosome matures toward the catalytic C complex.
The PPIase/cyclophilin domain of CWC27 is almost certainly catalytically inactive (a degenerate cyclophilin / pseudo-enzyme):
- [PMID:20676357 "Structural and biochemical characterization of the human cyclophilin family of peptidyl-prolyl isomerases", "No binding was detected for PPIL2, PPIL6, or SDCCAG-10, making these ... the first set of human cyclophilins that have been found incompetent to ligate cyclosporin"]. SDCCAG-10 = CWC27.
- [PMID:20676357 Table 1: "SDCCAG-10 ... no/no" for cyclosporin binding and tetrapeptide activity]. CWC27 showed neither cyclosporin binding nor isomerase activity against tetrapeptide substrates.
- Structural basis: CWC27 has a glutamic acid (Glu122) at the position equivalent to the catalytic Trp121 of PPIA; "glutamic acid in SDCCAG10" abrogates activity, and "Glutamic acid at this position seems to be incompatible with isomerase activity" PMID:20676357. The crystal structure (PDB 2HQ6, res 8-173) underlies this conclusion.
- UniProt curates CWC27 as "Probable inactive peptidyl-prolyl cis-trans isomerase" with a CAUTION: "Despite the fact that it belongs to the cyclophilin-type PPIase family, a report has shown that it has probably no peptidyl-prolyl cis-trans isomerase activity" [file:human/CWC27/CWC27-uniprot.txt].
- Consequently the GOA carries an explicit NOT enables peptidyl-prolyl cis-trans isomerase activity (GO:0003755, IDA, PMID:20676357). This negated annotation should be ACCEPTed; it correctly records the experimental finding of absent catalysis.
Implication for annotations:
- The InterPro2GO IEA transfer of protein folding (GO:0006457, GO_REF:0000002) is a family-level inference based on the cyclophilin/PPIase InterPro signature. Given that CWC27 is a demonstrated pseudo-PPIase with no isomerase/chaperone catalytic activity, this is an over-annotation (paralog/family transfer that ignores the loss of catalytic residues). MARK_AS_OVER_ANNOTATED.
CWC27 acts as a cyclophilin-fold scaffold within the spliceosome rather than as an enzyme. There is no well-fitting catalytic MF. A non-catalytic structural/scaffolding role within the spliceosome is best captured by the CC (spliceosome complex) and BP (mRNA splicing) annotations; no confident replacement MF term is proposed. Note CWC27 pairs with CWC22 to position the EJC core factor eIF4A3 for deposition (from the broader literature / biological hint); this is a scaffolding/adaptor role but not directly evidenced in the cached publications, so no new MF term is asserted.
The original 16 annotation source objects and stored GOA snapshot are retained, including the NOT PPIase qualifier. The genuine normal fetch added five WITH/FROM lists already present in the stored source; it did not replace the source inventory. A fresh 17-record QuickGO capture has one additional IEA catalytic-step-2 assertion (GO_REF:0000117), recorded separately without relabeling it NEW or replacing the old snapshot. The existing genuine Falcon report is preserved as secondary evidence, and the previous notes remain an exact prefix. Current conclusions below supersede categorical stage, catalytic-causality and no-folding statements in the earlier text.
The original 2002 study is stronger than an unspecified bulk-spliceosome observation. Its Table 1 lists NY-CO-10/O60529, a verified secondary accession of Q6UX04, with five unique C-complex peptides and none in H complex. The AG-to-GG 3′ splice-site mutation arrests after the first splicing step, and RNase H depletes earlier complexes. Affinity eluates contain 75–85% intermediate RNA; the separate gradient peak reaches 96%. Those percentages refer to RNA at different purification stages, not CWC27 purity or stoichiometry, and the target MS result is not specifically attributed to the gradient peak. This supports contextual step-2 membership and the associated splicing inference. Later structures do not prove the earlier target peptides were contamination. The reason for their different target occupancy remains unresolved. PMID:11991638(https://doi.org/10.1017/S1355838202021088)
The 2018 major spliceosome has U2/U5/U6 snRNPs and is not yet competent to branch: CWC27 is strong early, weaker mature and absent late Bact. GO:0071005 requires the all-five-snRNP precatalytic state, so that annotation is refined to GO:0005684 U2-type spliceosomal complex. The IC splicing inference remains biologically supported while its original component-term premise is scoped explicitly. In the 2024 six-state series CWC27 occupies Bact-I and is released in Bact-II; these source-specific classifications are not collapsed into one universal dissociation time. PMID:29360106(https://pmc.ncbi.nlm.nih.gov/articles/PMC5835773/), PMID:39068178(https://pmc.ncbi.nlm.nih.gov/articles/PMC11283556/)
The recovered 2021 minor-spliceosome original includes CWC27/NY-CO-10 in its 45-protein Bact model. MINX-U12 truncation 18 nucleotides beyond the branch-point sequence prevents PRP2 remodeling and traps a pre-branching state. GO:0071018, defined after first cleavage, is therefore refined to GO:0005689 U12-type spliceosomal complex. This is an actual source-state correction, not rejection of minor-spliceosome participation. Activated synonyms are not sufficient to infer the catalytic-step-1 GO terms either. PMID:33509932(https://doi.org/10.1126/science.abg0879)
The human CWC27 cyclophilin domain does not accelerate isomerization of the tested AAPF/AFPF/AGPF tetrapeptides and does not bind cyclosporin under the reported conditions, but a proline-containing peptide produces positive binding shifts. The NOT PPIase annotation is retained. An active-site comparison and a PPIA mutant do not constitute a CWC27 rescue proving one substitution is solely causal. Protein folding is broader than isomerase catalysis and includes noncatalytic assistance; neither the negative catalytic assays nor CWC22 stabilization settles that process. The IPR020892 protein-folding assertion is UNDECIDED. PMID:20676357(https://pmc.ncbi.nlm.nih.gov/articles/PMC2911226/)
Both official products reconstruct exactly: Q6UX04-1 is 472 residues, and Q6UX04-2 is 390 after VSP_030082/030083. The 2020 Methods explicitly use Q6UX04-2, which fails retention by recombinant CWC22-S with or without EIF4A3. Full-length CWC27 and its C-terminal region do bind CWC22. The structural peptide includes a CWC22-contacting helix; no direct CWC27–EIF4A3 contact is established. This is a measured product difference without a universal no-function claim for the short product. PMID:32329775(https://pmc.ncbi.nlm.nih.gov/articles/PMC7261170/)
The 2020 ternary reconstitution, structure and human-cell perturbations support a noncatalytic CWC22-associated contribution to splicing. Its proposed ordered landing platform is not the only established route. The 2026 human HEK293T study finds CWC27/CWC22–EIF4A3 coIP persisting when pladienolide B blocks pre-B formation and also detects CWC27 with EJC components. EIF4A3/EJC association can precede CWC27. Extra-spliceosomal recruitment and changed conformations are proposals; coIP does not establish direct CWC27–EIF4A3 binding. The one core therefore retains structural splicing participation, spliceosomal-complex membership and nucleoplasm, without inventing an intrinsic catalytic or adaptor MF or a new EJC process assertion. PMID:42635127(https://pmc.ncbi.nlm.nih.gov/articles/PMC13501132/)
Actual PTHR45625 has 1,072 nodes and MSA entries. The target leaf PTN002476198 matches the exact canonical sequence and descends from GO:0071013 IBD PTN000116297. Target self evidence is valid, with current human PPIL1/PPIL3 and CWC27 experimental descendants. The distinct on-path PPIase-loss record concerns another term. Current PPIL2 records and the unmatched current ARBA00026330 rule are snapshot differences, not grounds for erasing historical experimental evidence. The pending alternative PTHR45625-paint.tsv remains context only and was not adopted.
Human HPA records support nucleoplasm across antibodies and cell systems; assay XML and siRNA controls were inspected, not image pixels. Three Reactome roots yield seven actual target paths through human CWC27 entity 8867885 (residues 2–472). Input/output membership supports location, not CWC27 helicase activity, persistent B* occupancy, or a direct NS5 pair. No target activity occurs in the selected GO-CAM index. Current IntAct contains 36 complete records whose target backbones match canonical CWC27; no extra binding assertion is manufactured from that inventory.
Both author-hosted 2002/2021 original PDFs were recovered and selected operative sections inspected; their cached abstract files remain unchanged. The 2002 Table 1 was visually inspected. Selected main Methods/Results/Discussion and captions were read for 2010/2018/2020/2024/2026, with exact per-paper limits retained in the compact source index. The 2017 disease source was assessed from its complete abstract. No complete supplementary or figure-pixel reading is claimed where absent. New genuine publication caches were added only for 32329775 and 42635127 after exact ownership/preimage qualification. Detailed original responses and reproducible projections are retained in the immutable audit packet; the public index provides source IDs, public URLs and target-specific scope without a raw-data dump.
The core now records GO:1990935, splicing factor binding, grounded in the direct CWC27–CWC22 reconstitution and interface experiments of PMID:32329775(https://pmc.ncbi.nlm.nih.gov/articles/PMC7261170/). Reciprocal binding of purified recombinant proteins establishes CWC22 binding; EIF4A3 alone did not retain CWC27. The tested full-length 472-residue product and its C-terminal region support the interaction, whereas the tested 390-residue splice product does not. This specifies the molecular activity already described in the existing core, without assigning RNA chemistry, direct EIF4A3 binding, or a single obligatory EJC recruitment route. All 16 reviewed source assertions and their actions remain unchanged; no new annotation row is added.
The original 16 source assertions, both official product objects and all earlier notes are preserved. This appendix supersedes the preceding UNDECIDED protein-folding recommendation: the IPR020892 conserved-site mapping is now marked over-annotated because the demonstrated CWC27-specific activities do not establish folding. The signature description includes chaperone functions, and GO:0006457 is broader than catalysis; the negative isomerase result does not prove that every noncatalytic folding role is absent. The biological question and proposed folding experiment remain.
One NEW GO:1990935 IPI is added for tested full-length Q6UX04-1, based on reciprocal purified CWC22 binding in PMID:32329775. There was no existing binding-function source row. This is direct molecular-function evidence, independent of the derived review status. Q6UX04-2 remains negative in the reported recombinant-retention assay. The single core and its terms are retained. An adaptor assignment is not added: CWC27 stabilizes the CWC22-associated assembly, but the minimal structure and isolated retention assay do not establish direct CWC27–EIF4A3 contact. PMID:42635127 supplies positive cellular corroboration of the CWC27–CWC22 module and its EJC associations, while allowing more than one recruitment route.
The primary source guide gives public original links and exact page/section locators, including the two papers whose repository caches remain abstract-only. The authored index is navigation, so its repeated supporting-evidence pointers and VERIFIED self-rating are removed. Existing primary citation identities and all source-cache bytes remain unchanged. The nonspecific 2024 state-list excerpt is replaced by its target-release excerpt; the 2020 ternary-assembly excerpt is replaced by a product-specific excerpt, with a separate short direct-binding anchor. One 2026 inhibitor/coIP anchor is added. No new quotation from the 2010 or 2018 papers is added: earlier supplied notes already contain extensive inherited excerpts, retained as a historical prefix. The appendices and new source guide use paraphrase for these sources.
The description again states pseudoenzyme biology, PRP8 contact and transient activated-spliceosome occupancy, with exact assay limits in the evidence sections. Source-specific state series are not forced into one universal release time. A precise ontology question records the missing pre-branching Bact distinction without inventing an identifier or reusing an ambiguous activated synonym. The existing compact JSON index is retained to keep this change focused on evidence content.