6KC8

Crystal structure of WT Nme1Cas9 in complex with sgRNA and target DNA (ATATGATT PAM) in post-cleavage state

6KC8 の概要

• エントリーDOI: 10.2210/pdb6kc8/pdb
• 分子名称: CRISPR-associated endonuclease Cas9, sgRNA, DNA (5'-D(P*AP*AP*GP*TP*TP*AP*AP*AP*TP*AP*GP*CP*AP*GP*AP*GP*TP*GP*AP*CP*C)-3'), ... (6 entities in total)
• 機能のキーワード: crispr-cas9, nmecas9, nme1cas9, hydrolase, ternary complex, hydrolase-rna-dna complex, hydrolase/rna/dna
• 由来する生物種: Neisseria meningitidis 8013; 詳細
• タンパク質・核酸の鎖数: 5
• 化学式量合計: 181913.64

構造登録者

Sun, W.,Yang, J.,Cheng, Z.,Liu, C.,Wang, K.,Huang, X.,Wang, Y. (登録日: 2019-06-27, 公開日: 2019-11-06, 最終更新日: 2023-11-22)

主引用文献

Sun, W.,Yang, J.,Cheng, Z.,Amrani, N.,Liu, C.,Wang, K.,Ibraheim, R.,Edraki, A.,Huang, X.,Wang, M.,Wang, J.,Liu, L.,Sheng, G.,Yang, Y.,Lou, J.,Sontheimer, E.J.,Wang, Y.
Structures of Neisseria meningitidis Cas9 Complexes in Catalytically Poised and Anti-CRISPR-Inhibited States.
Mol.Cell, 76:938-, 2019

PubMed Abstract: High-resolution Cas9 structures have yet to reveal catalytic conformations due to HNH nuclease domain positioning away from the cleavage site. Nme1Cas9 and Nme2Cas9 are compact nucleases for in vivo genome editing. Here, we report structures of meningococcal Cas9 homologs in complex with sgRNA, dsDNA, or the AcrIIC3 anti-CRISPR protein. DNA-bound structures represent an early step of target recognition, a later HNH pre-catalytic state, the HNH catalytic state, and a cleaved-target-DNA-bound state. In the HNH catalytic state of Nme1Cas9, the active site is seen poised at the scissile phosphodiester linkage of the target strand, providing a high-resolution view of the active conformation. The HNH active conformation activates the RuvC domain. Our structures explain how Nme1Cas9 and Nme2Cas9 read distinct PAM sequences and how AcrIIC3 inhibits Nme1Cas9 activity. These structures provide insights into Cas9 domain rearrangements, guide-target engagement, cleavage mechanism, and anti-CRISPR inhibition, facilitating the optimization of these genome-editing platforms.

PubMed: 31668930
DOI: 10.1016/j.molcel.2019.09.025

実験手法

X-RAY DIFFRACTION (2.9 Å)