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Crystal Structure of Staphylococcus aureus Cas9

Author(s)
Nishimasu, Hiroshi; Ran, F. Ann; Kurabayashi, Arisa; Ishitani, Ryuichiro; Nureki, Osamu; Cong, Le; Yan, Winston Xia; Zetsche, Bernd; Li, Yinqing; Zhang, Feng; ... Show more Show less
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Abstract
Summary The RNA-guided DNA endonuclease Cas9 cleaves double-stranded DNA targets with a protospacer adjacent mot if (PAM) and complementarity to the guide RNA. Recently, we harnessed Staphylococcus aureus Cas9 (SaCas9), which is significantly smaller than Streptococcus pyogenes Cas9 (SpCas9), to facilitate efficient in vivo genome editing. Here, we report the crystal structures of SaCas9 in complex with a single guide RNA (sgRNA) and its double-stranded DNA targets, containing the 5′-TTGAAT-3′ PAM and the 5′-TTGGGT-3′ PAM, at 2.6 and 2.7 Å resolutions, respectively. The structures revealed the mechanism of the relaxed recognition of the 5′-NNGRRT-3′ PAM by SaCas9. A structural comparison of SaCas9 with SpCas9 highlighted both structural conservation and divergence, explaining their distinct PAM specificities and orthologous sgRNA recognition. Finally, we applied the structural information about this minimal Cas9 to rationally design compact transcriptional activators and inducible nucleases, to further expand the CRISPR-Cas9 genome editing toolbox.
Date issued
2015-08
URI
http://hdl.handle.net/1721.1/112724
Department
Institute for Medical Engineering and Science; Massachusetts Institute of Technology. Department of Biological Engineering; Massachusetts Institute of Technology. Department of Brain and Cognitive Sciences; McGovern Institute for Brain Research at MIT
Journal
Cell
Publisher
Elsevier
Citation
Nishimasu, Hiroshi et al. “Crystal Structure of Staphylococcus Aureus Cas9.” Cell 162, 5 (August 2015): 1113–1126 © 2015 Elsevier
Version: Author's final manuscript
ISSN
0092-8674
1097-4172

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