5B2P

Crystal structure of Francisella novicida Cas9 in complex with sgRNA and target DNA (TGA PAM)

5B2P の概要

• エントリーDOI: 10.2210/pdb5b2p/pdb
• 関連するPDBエントリー: 5B2O 5B2Q
• 分子名称: CRISPR-associated endonuclease Cas9, ACETATE ION, Guide RNA, ... (11 entities in total)
• 機能のキーワード: crispr-cas9, genome engineering, hydrolase-rna-dna complex, hydrolase/rna/dna
• 由来する生物種: Francisella tularensis subsp. novicida U112; 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 235532.22

構造登録者

Hirano, H.,Nishimasu, H.,Nakane, T.,Ishitani, R.,Nureki, O. (登録日: 2016-02-01, 公開日: 2016-03-02, 最終更新日: 2024-03-20)

主引用文献

Hirano, H.,Gootenberg, J.S.,Horii, T.,Abudayyeh, O.O.,Kimura, M.,Hsu, P.D.,Nakane, T.,Ishitani, R.,Hatada, I.,Zhang, F.,Nishimasu, H.,Nureki, O.
Structure and Engineering of Francisella novicida Cas9
Cell, 164:950-961, 2016

PubMed Abstract: The RNA-guided endonuclease Cas9 cleaves double-stranded DNA targets complementary to the guide RNA and has been applied to programmable genome editing. Cas9-mediated cleavage requires a protospacer adjacent motif (PAM) juxtaposed with the DNA target sequence, thus constricting the range of targetable sites. Here, we report the 1.7 Å resolution crystal structures of Cas9 from Francisella novicida (FnCas9), one of the largest Cas9 orthologs, in complex with a guide RNA and its PAM-containing DNA targets. A structural comparison of FnCas9 with other Cas9 orthologs revealed striking conserved and divergent features among distantly related CRISPR-Cas9 systems. We found that FnCas9 recognizes the 5'-NGG-3' PAM, and used the structural information to create a variant that can recognize the more relaxed 5'-YG-3' PAM. Furthermore, we demonstrated that the FnCas9-ribonucleoprotein complex can be microinjected into mouse zygotes to edit endogenous sites with the 5'-YG-3' PAM, thus expanding the target space of the CRISPR-Cas9 toolbox.

PubMed: 26875867
DOI: 10.1016/j.cell.2016.01.039

実験手法

X-RAY DIFFRACTION (1.7 Å)