5AXW

Crystal structure of Staphylococcus aureus Cas9 in complex with sgRNA and target DNA (TTGGGT PAM)

Summary for 5AXW

• Entry DOI: 10.2210/pdb5axw/pdb
• Related: 5CZZ
• Descriptor: CRISPR-associated endonuclease Cas9, RNA (73-MER), DNA (28-MER), ... (8 entities in total)
• Functional Keywords: crispr-cas9, genome engineering, hydrolase-rna-dna complex, hydrolase/rna/dna
• Biological source: Staphylococcus aureus subsp. aureus; More
• Total number of polymer chains: 4
• Total formula weight: 159140.07

Authors

Nishimasu, H.,Ishitani, R.,Nureki, O. (deposition date: 2015-08-01, release date: 2015-09-02, Last modification date: 2024-03-20)

Primary citation

Nishimasu, H.,Cong, L.,Yan, W.X.,Ran, F.A.,Zetsche, B.,Li, Y.,Kurabayashi, A.,Ishitani, R.,Zhang, F.,Nureki, O.
Crystal Structure of Staphylococcus aureus Cas9.
Cell, 162:1113-1126, 2015

PubMed Abstract: 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 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.

PubMed: 26317473
DOI: 10.1016/j.cell.2015.08.007

Experimental method

X-RAY DIFFRACTION (2.7 Å)