6AI6

Crystal structure of SpCas9-NG

6AI6 の概要

• エントリーDOI: 10.2210/pdb6ai6/pdb
• 分子名称: CRISPR-associated endonuclease Cas9/Csn1, RNA (81-MER), DNA (28-MER), ... (8 entities in total)
• 機能のキーワード: nuclease, hydrolase-rna-dna complex, hydrolase, hydrolase/rna/dna
• 由来する生物種: Streptococcus pyogenes serotype M1; 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 197022.33

構造登録者

Nishimasu, H.,Hirano, S.,Ishitani, R.,Nureki, O. (登録日: 2018-08-21, 公開日: 2018-10-31, 最終更新日: 2025-09-17)

主引用文献

Nishimasu, H.,Shi, X.,Ishiguro, S.,Gao, L.,Hirano, S.,Okazaki, S.,Noda, T.,Abudayyeh, O.O.,Gootenberg, J.S.,Mori, H.,Oura, S.,Holmes, B.,Tanaka, M.,Seki, M.,Hirano, H.,Aburatani, H.,Ishitani, R.,Ikawa, M.,Yachie, N.,Zhang, F.,Nureki, O.
Engineered CRISPR-Cas9 nuclease with expanded targeting space
Science, 361:1259-1262, 2018

PubMed Abstract: The RNA-guided endonuclease Cas9 cleaves its target DNA and is a powerful genome-editing tool. However, the widely used Cas9 enzyme (SpCas9) requires an NGG protospacer adjacent motif (PAM) for target recognition, thereby restricting the targetable genomic loci. Here, we report a rationally engineered SpCas9 variant (SpCas9-NG) that can recognize relaxed NG PAMs. The crystal structure revealed that the loss of the base-specific interaction with the third nucleobase is compensated by newly introduced non-base-specific interactions, thereby enabling the NG PAM recognition. We showed that SpCas9-NG induces indels at endogenous target sites bearing NG PAMs in human cells. Furthermore, we found that the fusion of SpCas9-NG and the activation-induced cytidine deaminase (AID) mediates the C-to-T conversion at target sites with NG PAMs in human cells.

PubMed: 30166441
DOI: 10.1126/science.aas9129

実験手法

X-RAY DIFFRACTION (2.7 Å)