UniProt entry is unreviewed (TrEMBL), 87 aa, SubName: Full=Anti-CRISPR protein
AcrIIA4 from EMBL and SubName: Full=Associated protein from PDB 5XBL. Taxon is
assigned to Listeria monocytogenes (NCBITaxon:1639) rather than to a phage, because
the gene sits in a prophage integrated in the L. monocytogenes chromosome.
The accession is the assayed protein. Rauch et al. used AcrIIA4- (AEO04689.1) as
the reference sequence for their phylogenetic reconstruction
PMID:28041849.
NCBI efetch of AEO04689.1 (hypothetical protein LMOG_02993, L. monocytogenes J0161)
returns
MNINDLIREIKNKDYTVKLSGTDSNSITQLIIRVNNDGNEYVISESENESIVEKFISAFKNGWNQEYEDEEEFYNDMQTITLKSELN,
which is 100% identical over all 87 residues to A0A247D711. The entry also carries
PDB 5XBL (AcrIIA4–SpyCas9–sgRNA, 3.05 Å) and 5XN4 (solution NMR), so the structural and
the genetic work are on the same sequence. Experimental evidence codes are therefore
appropriate on this accession, not similarity codes.
AcrIIA4 was one of four type II-A Cas9 inhibitors found by searching cas9-bearing
genomes for a CRISPR spacer coexisting with its own target
PMID:28041849.
Two of the four, AcrIIA2 and AcrIIA4, also inhibit the heterologous S. pyogenes Cas9
PMID:28041849,
and the authors attribute the breadth to a conserved target surface
PMID:28041849.
Prophage carriage is common, so this is a widespread counter-defence and not a curiosity
PMID:28041849.
The module (modules/anti_crispr_suppression.yaml, annoton acriia4_dna_mimic) assigns
AcrIIA4 to the "Type II Cas9 PAM-site occlusion and DNA mimicry" variant. The
assignment holds, and is if anything understated: two surfaces are occluded, not one.
Yang & Patel solved the AcrIIA4–SpyCas9–sgRNA crystal structure and found competitive
occupancy of both the PAM-reading surface and the RuvC pocket
PMID:28602637,
PMID:28602637.
The buried-area comparison is the quantitative statement of the competition
PMID:28602637,
and the β1–β2 loop reaches into the RuvC active site itself
PMID:28602637.
Dong et al., independently, state the mimicry explicitly
PMID:28448066
(abstract only in cache; no PMC record).
The physical basis of the mimicry is an acidic surface patch, characterised by NMR
PMID:29497118,
PMID:29497118.
The same paper notes convergence with the unrelated type I-F inhibitor AcrF2
PMID:29497118,
which is exactly the mechanism-not-family convergence the module is built around.
Binding is guide-dependent: the binding site on Cas9 only exists once sgRNA is loaded
PMID:28448066,
PMID:28448066,
and Yang & Patel devote a results section to it
PMID:28602637
— so AcrIIA4 engages a ribonucleoprotein, not apo Cas9.
Contrast with AcrIIC1, which also inhibits Cas9 but leaves DNA binding intact
PMID:28844692
(truncated in cache at this point) — the two mechanism classes in the module (part 1
recognition blockade vs part 2 nuclease neutralisation) are distinguished in the primary
literature itself.
There is no GO molecular-function term for mimicry-based competitive inhibition. QuickGO
searches for "molecular mimicry" return only GO:0140489 molecular template activity and
the generic regulator terms; nothing for a protein that imitates a nucleic acid to occupy
its binding site. GO:0043021 ribonucleoprotein complex binding is true of AcrIIA4
(binding requires sgRNA-loaded Cas9) but says nothing about mimicry, and
GO:0140721 nuclease inhibitor activity is true but flattens the mechanism to generic
inhibition. A new term nucleic acid mimicry-based competitive inhibitor activity is
proposed under GO:0140678 molecular function inhibitor activity; it would serve
AcrIIA4, AcrIIA2 (genes/9CAUD/acrIIA2) and AcrF2/gp30 (genes/BPD31/orf30) at minimum.
GOA (CACAO) holds one row: acts_upstream_of_or_within_positive_effect GO:0098672, IDA,
PMID:28041849. The term is right. The qualifier is not well chosen: AcrIIA4 does not
act upstream of the suppression process, it is the executing step — it binds Cas9
directly and the binding is the suppression. involved_in is the better-supported
qualifier, and acts_upstream_of_or_within_positive_effect additionally reads as though
AcrIIA4 positively regulated something, which inverts the sense for a reader skimming the
GAF. Same issue on A0A2D0TCG3 (genes/NEIME/acrIIC1). Flagged in review.reason;
the annotation itself is accepted.
AcrIIA4 is heavily used as a Cas9 "off-switch" reagent in human cells. That is a
laboratory application, not a biological function of the prophage gene, and is excluded
from the review's description and from all annotations.
PMID:28706995 supplies the missing kinetic boundary: 1:1 binding and inhibition before DNA recognition, but no effect on preformed Cas9-sgRNA-DNA complexes and no increase in DNA dissociation. The old suggestion that displacement remained untested was removed. The paper also supports near-stoichiometric inhibition, so universal inhibitor molar excess and rapid reversibility were removed from the description and shared NTR definition. Accession identity is exact to AEO04689.1; the record remains bacterial because its sequence originates in a resident prophage.
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-LISMN-acrIIA4.log. These are manual research notes, not provider-generated research.