7L48

Cryo-EM structure of a CRISPR-Cas12f Binary Complex

7L48 の概要

• エントリーDOI: 10.2210/pdb7l48/pdb
• EMDBエントリー: 23157
• 分子名称: Cas12f, sgRNA, ZINC ION (3 entities in total)
• 機能のキーワード: crispr cas, rna binding protein, rna binding protein-rna complex, rna binding protein/rna
• 由来する生物種: unidentified; 詳細
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 196059.47

構造登録者

Chang, L.,Li, Z. (登録日: 2020-12-18, 公開日: 2021-06-02, 最終更新日: 2024-03-06)

主引用文献

Xiao, R.,Li, Z.,Wang, S.,Han, R.,Chang, L.
Structural basis for substrate recognition and cleavage by the dimerization-dependent CRISPR-Cas12f nuclease.
Nucleic Acids Res., 49:4120-4128, 2021

PubMed Abstract: Cas12f, also known as Cas14, is an exceptionally small type V-F CRISPR-Cas nuclease that is roughly half the size of comparable nucleases of this type. To reveal the mechanisms underlying substrate recognition and cleavage, we determined the cryo-EM structures of the Cas12f-sgRNA-target DNA and Cas12f-sgRNA complexes at 3.1 and 3.9 Å, respectively. An asymmetric Cas12f dimer is bound to one sgRNA for recognition and cleavage of dsDNA substrate with a T-rich PAM sequence. Despite its dimerization, Cas12f adopts a conserved activation mechanism among the type V nucleases which requires coordinated conformational changes induced by the formation of the crRNA-target DNA heteroduplex, including the close-to-open transition in the lid motif of the RuvC domain. Only one RuvC domain in the Cas12f dimer is activated by substrate recognition, and the substrate bound to the activated RuvC domain is captured in the structure. Structure-assisted truncated sgRNA, which is less than half the length of the original sgRNA, is still active for target DNA cleavage. Our results expand our understanding of the diverse type V CRISPR-Cas nucleases and facilitate potential genome editing applications using the miniature Cas12f.

PubMed: 33764415
DOI: 10.1093/nar/gkab179

実験手法

ELECTRON MICROSCOPY (3.9 Å)