期刊
CELL
卷 162, 期 5, 页码 1113-1126出版社
CELL PRESS
DOI: 10.1016/j.cell.2015.08.007
关键词
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资金
- JST
- JSPS KAKENHI [26291010, 15H01463]
- Ministry of Education, Culture, Sports, Science and Technology
- National Institute of General Medical Sciences [T32GM007753]
- Paul and Daisy Soros Fellowship
- NIH [1DP1-MH100706]
- Keck Foundation
- McKnight Foundation
- Poitras Foundation
- Merkin Foundation
- Vallee Foundation
- Damon Runyon Foundation
- Searle Scholars Foundation
- Klingenstein Foundation
- Simons Foundation
- Japan Agency for Medical Research and Development, AMED
- Council for Science, Technology and Innovation (CSTI)
- Cross-ministerial Strategic Innovation Promotion Program (SIP)
- Technologies for creating next-generation agriculture, forestry and fisheries'' (Bio-oriented Technology Research Advancement Institution, NARO)
- PRESTO
- Grants-in-Aid for Scientific Research [26291010, 15H01463] Funding Source: KAKEN
The RNA-guided DNA endonuclease Cas9 cleaves double-stranded DNA targets with a protospacer adjacent motif (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 angstrom 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.
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