Summary information and primary citation

PDB-id
6ai6; DSSR-derived features in text and JSON formats
Class
hydrolase-RNA-DNA
Method
X-ray (2.7 Å)
Summary
Crystal structure of spcas9-ng
Reference
Nishimasu H, Shi X, Ishiguro S, Gao L, Hirano S, Okazaki S, Noda T, Abudayyeh OO, Gootenberg JS, Mori H, Oura S, Holmes B, Tanaka M, Seki M, Hirano H, Aburatani H, Ishitani R, Ikawa M, Yachie N, Zhang F, Nureki O (2018): "Engineered CRISPR-Cas9 nuclease with expanded targeting space." Science, 361, 1259-1262. doi: 10.1126/science.aas9129.
Abstract
The RNA-guided endonuclease Cas9 cleaves its target DNA and is a powerful genome-editing tool. However, the widely used Streptococcus pyogenes 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.

Cartoon-block schematics in six views (download the tarball)

PyMOL session file Download PDB file View in 3Dmol.js