Structures of a CRISPR-Cas9 R-loop complex primed for DNA cleavage

被引：467

作者：

Jiang, Fuguo ^{[1
]}

Taylor, David W. ^{[1
,2
]}

Chen, Janice S. ^{[1
]}

Kornfeld, Jack E. ^{[3
]}

Zhou, Kaihong ^{[3
]}

Thompson, Aubri J. ^{[4
]}

Nogales, Eva ^{[1
,2
,3
,5
]}

Doudna, Jennifer A. ^{[1
,2
,3
,4
,5
,6
]}

机构：

[1] Univ Calif Berkeley, Dept Mol & Cell Biol, 229 Stanley Hall, Berkeley, CA 94720 USA

[2] Univ Calif Berkeley, Calif Inst Quantitat Biosci, Berkeley, CA 94720 USA

[3] Univ Calif Berkeley, Howard Hughes Med Inst, Berkeley, CA 94720 USA

[4] Univ Calif Berkeley, Dept Chem, Berkeley, CA 94720 USA

[5] Univ Calif Berkeley, Lawrence Berkeley Natl Lab, Div Life Sci, Berkeley, CA 94720 USA

[6] Univ Calif Berkeley, Lawrence Berkeley Natl Lab, Phys Biosci Div, Berkeley, CA 94720 USA

来源：

SCIENCE | 2016年 / 351卷 / 6275期

基金：

美国国家科学基金会;

关键词：

CAS SYSTEMS; TARGET DNA; ADAPTIVE IMMUNITY; CRYSTAL-STRUCTURE; RNA; ENDONUCLEASE; RECOGNITION; BACTERIA; CASCADE; PROKARYOTES;

D O I：

10.1126/science.aad8282

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

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 degrees, providing the structural distortion needed for R-loop formation.

引用

页码：867 / 871

页数：5