Unreviewed TrEMBL entry, 174 aa, SubName: Full=AcrVA1 {ECO:0000313|PDB:6NMD} — the name
comes from a PDB deposition. GN ORFNames=AAX09_07405, a Moraxella bovoculi locus tag.
PE 1: Evidence at protein level. CDD cd22787 AcrVA1. Seeded taxon was wrong
(NCBITaxon:9, which is Buchnera aphidicola); UniProt gives OX NCBI_TaxID=386891
(Moraxella bovoculi) and the review is corrected to that. No GOA annotations at all.
PDB; 6NMD; EM; 3.49 A; B=5-174 is the LbCas12a–crRNA–AcrVA1 complex; RCSB confirms its
primary citation is PMID:31155345 and its structure title is "cryo-EM Structure of the
LbCas12a-crRNA-AcrVA1 complex". So the structural work is on this accession.
For PMID:30936531 (Knott et al.), the methods say only that the AcrVA1 expression plasmid
was "generated from a custom pET-based expression vector as described previously18"
(ref 18 = Marino et al. 2018), with no accession. Both papers work on the Moraxella
bovoculi AcrVA1, which is the canonical one, but I cannot verify sequence identity from
the cached text. The structural/binding annotations are therefore grounded on
PMID:31155345 (direct, via 6NMD), and PMID:30936531 is used for the enzymology with that
caveat recorded.
The module (annoton acrva1_crrna_nuclease) places AcrVA1 in the
crrna_cleavage_variant of the enzymatic-subversion part, and asserts
required_function and function = GO:0004521 RNA endonuclease activity, with substrate
"Cas12a-bound crRNA guide". The mechanism class is correct. The attribution of the
catalytic centre to AcrVA1 is not established, and the module asserts it without
qualification.
What is established:
What is not established — the catalyst. The activity requires both AcrVA1 and
Cas12a-crRNA together; AcrVA1 alone does nothing to RNA
PMID:30936531,
and Knott et al. say explicitly that they could not assign the catalytic centre
PMID:30936531.
They also report that mutating either known Cas12a nuclease centre (RuvC, and the
pre-crRNA processing nuclease) does not abolish the truncation, so the obvious alternative
candidates are excluded without the AcrVA1 option being confirmed. The independent
structural paper states the active reading without qualification
PMID:31155345
but the abstract alone does not identify catalytic residues. The later publisher-highlight inspection recorded below reports a candidate helix-alpha2 active site; the earlier categorical absence claim is superseded.
Curation decision. GO:0004521 RNA endonuclease activity is asserted, because two
independent groups report Cas12a-dependent endoribonucleolytic cleavage with multiple
turnover and the balance of the published interpretation is that AcrVA1 is the nuclease.
But it is asserted with the caveat in review.reason and with an explicit BIOLOGY
knowledge gap, because the alternative — that AcrVA1 allosterically licenses a latent
Cas12a ribonuclease activity — has not been excluded, and if it were true the term would
belong on Cas12a and AcrVA1 would instead be an enzyme activator. This is the point at
which the module should be softened: required_function: GO:0004521 makes the
unestablished attribution a membership criterion for the family, which is stronger than
the evidence.
GO:0043021 ribonucleoprotein complex binding is also asserted, and is on firmer ground
than the catalytic term: the cryo-EM structure is of AcrVA1 bound to the Cas12a-crRNA
ribonucleoprotein, and every activity AcrVA1 has is conditional on that complex existing.
AcrVA1 inserts into the PAM-binding pocket, which is superficially the AcrIIA4/AcrIIA2
mechanism. It is not annotated as a nucleic acid mimic here, because neither paper
describes an acidic pseudo-DNA surface or competition with the duplex; the PAM-pocket
insertion is reported as the positioning that puts AcrVA1 next to the crRNA spacer it then
cleaves. Assigning mimicry would be reading the AcrIIA4 story onto a different protein.
The module's placement of AcrVA1 in part 3 (enzymatic) rather than part 1 (recognition
blockade) is the right call, even though AcrVA1 does also block dsDNA binding.
Unreviewed entry, PDB-derived name, zero GOA annotations, for a protein whose mechanism
was called "a mode of inhibition not previously observed for any anti-CRISPR protein".
Same curation gap class as L7P7R7 and A0A2D0TCG3.
PMID:30936531 establishes spacer cleavage, multiple turnover and release of the 3-prime fragment, not loss of the complete guide or permanent impossibility of reprogramming. PMID:31155345 (cached abstract; publisher highlights additionally identify a helix-alpha2 RNase active site) is later independent evidence, so the old claim that no candidate catalytic site had been reported was removed. The direct structural accession link is PDB 6NMD chain B, residues 5-174. The prior reviewer-authored host-cell-cytoplasm proposal was corrected to cytoplasm because the source record is bacterial and the native element is unresolved. No GOA assertion was changed.
Existing GOA assertions were preserved. Earlier reviewer-authored NEW proposals were reassessed independently. Primary references were obtained through the normal publication cache. A normal Falcon attempt with perplexity-lite fallback failed; logs are in .cache/module-audit-logs/anti-crispr-deep-9GAMM-acrVA1.log. These are manual research notes, not provider-generated research.
Restored the catalytic-entity caveat in the GO:0004521 IC reason. PMID:30936531 directly states that mutating Cas12a RuvC or pre-crRNA-processing centres did not prevent truncation but does not assign the catalytic component; a latent Cas12a RNase licensed by AcrVA1 would make AcrVA1 the activator, not the nuclease. PMID:31155345 publisher highlights explicitly identify an RNase active site in helix alpha2 (official ScienceDirect page inspected again), so the obsolete claim that no candidate site exists is not restored. Normal PMID refresh still supplied abstract only and normal publisher URL fetching returned403. The IC proposal remains conditional, without a fabricated full-text quote or conversion to IDA.
The second core now uses contributes_to_molecular_function: GO:0004521 for the demonstrable AcrVA1–Cas12a-crRNA assembly activity. This makes the conservative catalytic-component boundary explicit in the structured summary while preserving the original IC proposal, its action and qualifier, and all source quotes. It does not assert that AcrVA1 lacks catalytic residues or cannot be activated by complex assembly.
Re-read the complete cached PMID:30936531 Results and Discussion: multiple turnover and known Cas12a nuclease-site mutagenesis establish the reaction and exclude those tested sites, but the authors explicitly do not assign the catalytic center. Re-read the PMID:31155345 abstract, which attributes cleavage to AcrVA1. The earlier inspection of publisher highlights reported a helix-alpha2 RNase site; the current publisher retry returned HTTP 403 and a targeted search did not recover the full paper. No new claim to have inspected its residue-mutant experiments is made, and its published interpretation remains visible. The structured contribution is a conservative statement of the shared experimental result, not a rejection of the later paper.