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Anti-CRISPR type V subtype A2 and similar proteins AcrVA2 is an anti-CRISPR (Acr) protein that was discovered via STSS (self-targeting spacers search) with further functional TXTL (cell-free transcription-translation) assay. The bioinformatic STSS method finds self-targeting CRISPR spacers in genomic DNA, predicts the type of CRISPR system involved, and obtains information about the targeted sequence. AcrVA2 is found distributed throughout multiple classes of bacteria with orthologs present in Lachnospiraceae and Leptospira. Distant orthologs of AcrVA2 are also present on plasmids and conjugative elements in Escherichia coli, such as protein FinQ, a transcriptional inhibitor of the F plasmid transfer gene. AcrVA2 has been shown to inhibit a CRISPR-associated nuclease Cas12a ortholog MbCas12a in Pseudomonas aeruginosa strains. Cas12a (previously called Cpf1) is an effector protein for the class 2, subtype V-A CRISPR-Cas system that has high efficiency in genome editing. Cas12a has a single nuclease domain (RuvC) that is activated upon binding of the crRNA targeting sequence (or spacer) to a complementary single-stranded DNA (ssDNA) or double-stranded DNA (dsDNA) target molecule; Cas12a also has non-specific ssDNA cleavage activity, known as promiscuous cleavage of collateral ssDNAs or trans-cleavage. In addition to gene editing, CRISPR-Cas12a has been re-purposed, in high eukaryotes, to carry out transcription regulation via a nuclease-dead Cas12a (dCas12a). Thus, AcrVAs have been adopted to regulate dCas12a activity. CRISPR-Cas immune systems are used by certain prokaryotes and archaea to resist the invasion of foreign nucleic acids such as phages or plasmids. Acr proteins are small proteins which are the natural inhibitors for CRISPR-Cas systems; encoded on bacterial and archaeal viruses, they allow the virus to evade host CRISPR-Cas systems. The CRISPR-Cas-mediated adaptive immune response can be divided into three steps, including the acquisition of spacer derived from invading nucleic acids, crRNA processing, and target degradation. Theoretically, Acr proteins could suppress any step to disrupt the CRISPR-Cas system. Acr proteins are diverse with no common sequence or structural motif, and they inhibit a wide range of CRISPR-Cas systems with various inhibition mechanisms. CRISPR-Cas systems are divided into two classes (1 and 2) and six types (class 1: types I, III and IV; class 2: types II, V and VI). The distinguishing feature of types II, V and VI is a single, large, multi-domain effector protein, such as Cas12a in type V subtype A. Type V systems appear to have evolved, on multiple independent occasions, from TnpB proteins (predicted RuvC-like nucleases, encoded by IS605-like transposons) yielding a large pool of type V variants, many of which may become separate subtypes. Acr families are named for their type and subtype which are numbered sequentially as they are discovered. |
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