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Mol Cell. 2019 Feb 7;73(3):611-620.e3. doi: 10.1016/j.molcel.2018.11.011. Epub 2018 Dec 31.

Phage AcrIIA2 DNA Mimicry: Structural Basis of the CRISPR and Anti-CRISPR Arms Race.

Author information

1
Key Laboratory of RNA Biology, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
2
Key Laboratory of RNA Biology, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China; State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China. Electronic address: ylwang@ibp.ac.cn.

Abstract

CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR-associated proteins) systems provide prokaryotic cells with adaptive immunity against invading bacteriophages. Bacteriophages counteract bacterial responses by encoding anti-CRISPR inhibitor proteins (Acr). However, the structural basis for their inhibitory actions remains largely unknown. Here, we report the crystal structure of the AcrIIA2-SpyCas9-sgRNA (single-guide RNA) complex at 3.3 Å resolution. We show that AcrIIA2 binds SpyCas9 at a position similar to the target DNA binding region. More specifically, AcrIIA2 interacts with the protospacer adjacent motif (PAM) recognition residues of Cas9, preventing target double-stranded DNA (dsDNA) detection. Thus, phage-encoded AcrIIA2 appears to act as a DNA mimic that blocks subsequent dsDNA binding by virtue of its highly acidic residues, disabling bacterial Cas9 by competing with target dsDNA binding with a binding motif distinct from AcrIIA4. Our study provides a more detailed mechanistic understanding of AcrIIA2-mediated inhibition of SpyCas9, the most widely used genome-editing tool, opening new avenues for improved regulatory precision during genome editing.

KEYWORDS:

AcrIIA2; SpyCas9; anti-CRISPR protein; sgRNA

PMID:
30606466
DOI:
10.1016/j.molcel.2018.11.011
[Indexed for MEDLINE]

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