Structures of Cas9 Endonucleases Reveal RNA-Mediated Conformational Activation
Top Cited Papers
- 14 March 2014
- journal article
- research article
- Published by American Association for the Advancement of Science (AAAS) in Science
- Vol. 343 (6176), 1247997
- https://doi.org/10.1126/science.1247997
Abstract
Type II CRISPR (clustered regularly interspaced short palindromic repeats)–Cas (CRISPR-associated) systems use an RNA-guided DNA endonuclease, Cas9, to generate double-strand breaks in invasive DNA during an adaptive bacterial immune response. Cas9 has been harnessed as a powerful tool for genome editing and gene regulation in many eukaryotic organisms. We report 2.6 and 2.2 angstrom resolution crystal structures of two major Cas9 enzyme subtypes, revealing the structural core shared by all Cas9 family members. The architectures of Cas9 enzymes define nucleic acid binding clefts, and single-particle electron microscopy reconstructions show that the two structural lobes harboring these clefts undergo guide RNA–induced reorientation to form a central channel where DNA substrates are bound. The observation that extensive structural rearrangements occur before target DNA duplex binding implicates guide RNA loading as a key step in Cas9 activation.Keywords
This publication has 87 references indexed in Scilit:
- CRISPR-Mediated Adaptive Immune Systems in Bacteria and ArchaeaAnnual Review of Biochemistry, 2013
- crRNA and tracrRNA guide Cas9-mediated DNA interference inStreptococcus thermophilusRNA Biology, 2013
- Cas9–crRNA ribonucleoprotein complex mediates specific DNA cleavage for adaptive immunity in bacteriaProceedings of the National Academy of Sciences of the United States of America, 2012
- A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial ImmunityScience, 2012
- RNA-guided genetic silencing systems in bacteria and archaeaNature, 2012
- CRISPR-based adaptive immune systemsCurrent Opinion in Microbiology, 2011
- Evolution and classification of the CRISPR–Cas systemsNature Reviews Microbiology, 2011
- Clustered regularly interspaced short palindromic repeats (CRISPRs): the hallmark of an ingenious antiviral defense mechanism in prokaryotesBiological Chemistry, 2011
- Small CRISPR RNAs Guide Antiviral Defense in ProkaryotesScience, 2008
- CRISPR Provides Acquired Resistance Against Viruses in ProkaryotesScience, 2007