A high‐resolution (1.2 Å) crystal structure of the anti‐CRISPR protein AcrIF9

Abstract

Acr, anti-CRISPR proteins EM, electron microscopy LB, lysogeny broth IPTG, isopropyl β-D-1-thiogalactopyranoside FPLC, fast protein liquid chromatography MR, molecular replacement SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis SEC, size-exclusion chromatography MALS, multi-angle light scattering RMSD, root mean square deviation

Mechanisms regulating the CRISPR-Cas systems

“…Target DNA binding AcrIF1 binds to the Csy complex (to Cas7 protein; Pawluk et al, 2016;Bondy-Denomy et al, 2018) AcrIF2 binds to the Cascade complex (to Cas8 and Cas7 proteins; Chowdhury et al, 2017) AcrIF4 binds to the Cascade complex but the exact binding locations is unknown (Bondy-Denomy et al, 2015;Pawluk et al, 2016) AcrIF9 binds to the Cascade complex and induces system to bind dsDNA independent of sequence complementarity or PAM (Hirschi et al, 2020;Kim et al, 2020;Lu et al, 2021) AcrIF10 binds to the Cascade complex (to Cas8 and Cas5 proteins; Pawluk et al, 2016;Guo et al, 2017) AcrID1 binds to the Cascade complex (to Cas10; He et al, 2018) AcrIIA2 binds to the Cas9 endonuclease but the exact binding locations is unknown (Rauch et al, 2017;Bondy-Denomy et al, 2018) AcrIIA4 mimics double-stranded DNA, binds to the Cas9-sgRNA complex (Shin et al, 2017;Yang and Patel, 2017) AcrIIA6 acts as an allosteric inhibitor and induces Cas9 dimerization (Hynes et al, 2018;Fuchsbauer et al, 2019;Zhang et al, 2019) AcrVA1 cuts off the 3′ end of the crRNA (Knott et al, 2019;Zhang et al, 2019) AcrVA4 dimerizes the Cas12a-crRNA complex (Knott et al, 2019) AcrVA5 acylates the Cas12 lysine residue that interacts with the PAM motif (Watters et al, 2018;Dong et al, 2019) AcrIIC3 dimerizes the Cas9 endonuclease (Sun et al, 2019) AcrIIC4 interacts with Cas9 but the exact mechanism is unknown (Lee et al, 2018) AcrIIC5 interacts with Cas9 but the exact mechanism is unknown (Lee et al, 2018) Target DNA cleavage AcrIIC2 interactions with the positively charged bridge helix of Cas9 endonuclease, thereby preventing sgRNA loading (Thavalingam et al, 2019;Zhu et al, 2019) AcrIIC1 binds to the HNH domain of the Cas9 endonuclease (Zhu et al, 2019) AcrIIA11 interacts with Cas9 and dsDNA but the exact mechanism is unknown (does not prevent target recognition;…”

Molecular basis of transcriptional repression of anti-CRISPR by anti-CRISPR-associated 2

“…To counteract the CRISPR-Cas systems of bacteria and archaea that confer resistance to foreign genetic material from invaders, the evolution of phages has resulted in the development of multiple anti-CRISPR (Acr) proteins that can inhibit the host CRISPR-Cas machinery via different strategies (Borges et al, 2017;Bondy-Denomy, 2018;Zhu et al, 2018;Kim et al, 2020;Lee et al, 2020). Identifying acr genes is difficult as they are highly divergent in sequence.…”