title: Structures of Cas9 endonucleases reveal RNA-mediated conformational activation.
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.
authors: Department of Biochemistry, University of Zurich, CH-8057 Zurich, Switzerland.
authors: Martin
authors: M
authors: Jinek
authors: Fuguo
authors: F
authors: Jiang
authors: David W
authors: DW
authors: Taylor
authors: Samuel H
authors: SH
authors: Sternberg
authors: Emine
authors: E
authors: Kaya
authors: Enbo
authors: E
authors: Ma
authors: Carolin
authors: C
authors: Anders
authors: Michael
authors: M
authors: Hauer
authors: Kaihong
authors: K
authors: Zhou
authors: Steven
authors: S
authors: Lin
authors: Matias
authors: M
authors: Kaplan
authors: Anthony T
authors: AT
authors: Iavarone
authors: Emmanuelle
authors: E
authors: Charpentier
authors: Eva
authors: E
authors: Nogales
authors: Jennifer A
authors: JA
authors: Doudna
mesh_terms: D000190:Actinomyces / Q000201:enzymology*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry*; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D018360:Crystallography, X-Ray; D053837:DNA Cleavage; D004720:Endonucleases / Q000737:chemistry*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry*; D013297:Streptococcus pyogenes / Q000201:enzymology*
chemical_list: D001426:Bacterial Proteins; D012313:RNA; D004720:Endonucleases