EMD-3277
Structures of a CRISPR-Cas9 R-loop complex primed for DNA cleavage
EMD-3277
Single-particle6.0 Å
Deposition: 10/12/2015Map released: 27/01/2016
Last modified: 16/03/2016
Sample Organism:
Streptococcus pyogenes
Sample: Cas9 bound to single guide-RNA and 40-bp target DNA
Deposition Authors: Jiang F, Taylor DW
,
Chen JS ,
Kornfeld JE,
Zhou K,
Thompson AJ,
Nogales E,
Doudna JA
Sample: Cas9 bound to single guide-RNA and 40-bp target DNA
Deposition Authors: Jiang F, Taylor DW
,
Chen JS ,
Kornfeld JE,
Zhou K,
Thompson AJ,
Nogales E,
Doudna JA
Structures of a CRISPR-Cas9 R-loop complex primed for DNA cleavage.
Jiang F,
Taylor DW ,
Chen JS ,
Kornfeld JE,
Zhou K,
Thompson AJ,
Nogales E,
Doudna JA
(2016) Science , 351 , 867 - 871
,
Chen JS ,
Kornfeld JE,
Zhou K,
Thompson AJ,
Nogales E,
Doudna JA
(2016) Science , 351 , 867 - 871
Abstract:
Bacterial adaptive immunity and genome engineering involving the CRISPR (clustered regularly interspaced short palindromic repeats)-associated (Cas) protein Cas9 begin with RNA-guided DNA unwinding to form an RNA-DNA hybrid and a displaced DNA strand inside the protein. The role of this R-loop structure in positioning each DNA strand for cleavage by the two Cas9 nuclease domains is unknown. We determine molecular structures of the catalytically active Streptococcus pyogenes Cas9 R-loop that show the displaced DNA strand located near the RuvC nuclease domain active site. These protein-DNA interactions, in turn, position the HNH nuclease domain adjacent to the target DNA strand cleavage site in a conformation essential for concerted DNA cutting. Cas9 bends the DNA helix by 30°, providing the structural distortion needed for R-loop formation.
Bacterial adaptive immunity and genome engineering involving the CRISPR (clustered regularly interspaced short palindromic repeats)-associated (Cas) protein Cas9 begin with RNA-guided DNA unwinding to form an RNA-DNA hybrid and a displaced DNA strand inside the protein. The role of this R-loop structure in positioning each DNA strand for cleavage by the two Cas9 nuclease domains is unknown. We determine molecular structures of the catalytically active Streptococcus pyogenes Cas9 R-loop that show the displaced DNA strand located near the RuvC nuclease domain active site. These protein-DNA interactions, in turn, position the HNH nuclease domain adjacent to the target DNA strand cleavage site in a conformation essential for concerted DNA cutting. Cas9 bends the DNA helix by 30°, providing the structural distortion needed for R-loop formation.