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TitleTemperature-Responsive Competitive Inhibition of CRISPR-Cas9.
Journal, issue, pagesMol Cell, Vol. 73, Issue 3, Page 601-610.e5, Year 2019
Publish dateFeb 7, 2019
AuthorsFuguo Jiang / Jun-Jie Liu / Beatriz A Osuna / Michael Xu / Joel D Berry / Benjamin J Rauch / Eva Nogales / Joseph Bondy-Denomy / Jennifer A Doudna /
PubMed AbstractCRISPR-Cas immune systems utilize RNA-guided nucleases to protect bacteria from bacteriophage infection. Bacteriophages have in turn evolved inhibitory "anti-CRISPR" (Acr) proteins, including six ...CRISPR-Cas immune systems utilize RNA-guided nucleases to protect bacteria from bacteriophage infection. Bacteriophages have in turn evolved inhibitory "anti-CRISPR" (Acr) proteins, including six inhibitors (AcrIIA1-AcrIIA6) that can block DNA cutting and genome editing by type II-A CRISPR-Cas9 enzymes. We show here that AcrIIA2 and its more potent homolog, AcrIIA2b, prevent Cas9 binding to DNA by occluding protein residues required for DNA binding. Cryo-EM-determined structures of AcrIIA2 or AcrIIA2b bound to S. pyogenes Cas9 reveal a mode of competitive inhibition of DNA binding that is distinct from other known Acrs. Differences in the temperature dependence of Cas9 inhibition by AcrIIA2 and AcrIIA2b arise from differences in both inhibitor structure and the local inhibitor-binding environment on Cas9. These findings expand the natural toolbox for regulating CRISPR-Cas9 genome editing temporally, spatially, and conditionally.
External linksMol Cell / PubMed:30595438 / PubMed Central
MethodsEM (single particle)
Resolution3.4 - 3.9 Å
Structure data

9066

6mcb

EMDB-9066, PDB-6mcb:
CryoEM structure of AcrIIA2 in complex with CRISPR-Cas9
Method: EM (single particle) / Resolution: 3.4 Å

9067

6mcc

EMDB-9067, PDB-6mcc:
CryoEM structure of AcrIIA2 homolog in complex with CRISPR-Cas9
Method: EM (single particle) / Resolution: 3.9 Å

Source
  • streptococcus pyogenes m1 gas (bacteria)
  • listeria phage lp-101 (virus)
KeywordsHYDROLASE/RNA/VIRAL PROTEIN / CRISPR-Cas / Cas9 / Cas9 inhibitors / anti-CRISPR / AcrIIA2 / bacteriophage / gene editing / HYDROLASE-RNA-VIRAL PROTEIN complex

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