CRISPR-Cas are bacterial and archaeal defense systems that provide resistance against phages and other invasive genomic elements. Short DNA fragments of extracellular origin are inserted within repeated sequences of the CRISPR locus. The CRISPR transcript is matured into small crRNA molecules, which, together with Cas proteins, assemble an effector complex destined to detect and destroy foreign nucleic acids [1]. The bacteriophages evolved small anti-CRISPR (Acr) proteins, which block CRISPR-Cas protection [2]. To date, over 100 Acr families have been discovered, but only a fraction of them have been studied in detail [3]. Recently, Acr proteins have been utilized to regulate CRISPR-Cas editing tools, demonstrating their immense practical potential.
Our research object is the Acr proteins inhibiting the type I-F CRISPR-Cas system from Aggregatibacter actinomycetemcomitans. The system encodes a ribonucleoprotein complex, termed Cascade, which binds foreign DNA and then recruits the Cas2/3 nuclease to destroy the invader [4]. We show Acr proteins blocking the CRISPR-Cas system in bacterial cells. We employ biochemical assays for the analysis of their molecular mechanisms. Our findings provide insight into the inhibitory mechanism of these previously uncharacterised proteins.