9CJH

Cas12a:gRNA:DNA (Acidaminococcus sp.) with 0 RNA:DNA base pairs, structure 1

9CJH の概要

• エントリーDOI: 10.2210/pdb9cjh/pdb
• 関連するPDBエントリー: 9CJI 9CJJ
• EMDBエントリー: 45631
• 分子名称: guide RNA (25-MER), CRISPR-associated endonuclease Cas12a, Non-target DNA strand, ... (4 entities in total)
• 機能のキーワード: dnase, complex, ribonucleoprotein, genome editor, hydrolase-rna complex, hydrolase, hydrolase-rna-dna complex, hydrolase/rna/dna
• 由来する生物種: Acidaminococcus sp. BV3L6; 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 184223.62

構造登録者

Soczek, K.M.,Doudna, J.A. (登録日: 2024-07-06, 公開日: 2025-01-08, 最終更新日: 2025-05-28)

主引用文献

Soczek, K.M.,Cofsky, J.C.,Tuck, O.T.,Shi, H.,Doudna, J.A.
CRISPR-Cas12a bends DNA to destabilize base pairs during target interrogation.
Nucleic Acids Res., 53:-, 2025

PubMed Abstract: RNA-guided endonucleases are involved in processes ranging from adaptive immunity to site-specific transposition and have revolutionized genome editing. CRISPR-Cas9, -Cas12 and related proteins use guide RNAs to recognize ∼20-nucleotide target sites within genomic DNA by mechanisms that are not yet fully understood. We used structural and biochemical methods to assess early steps in DNA recognition by Cas12a protein-guide RNA complexes. We show here that Cas12a initiates DNA target recognition by bending DNA to induce transient nucleotide flipping that exposes nucleobases for DNA-RNA hybridization. Cryo-EM structural analysis of a trapped Cas12a-RNA-DNA surveillance complex and fluorescence-based conformational probing show that Cas12a-induced DNA helix destabilization enables target discovery and engagement. This mechanism of initial DNA interrogation resembles that of CRISPR-Cas9 despite distinct evolutionary origins and different RNA-DNA hybridization directionality of these enzyme families. Our findings support a model in which RNA-mediated DNA interference begins with local helix distortion by transient CRISPR-Cas protein binding.

PubMed: 39698811
DOI: 10.1093/nar/gkae1192

実験手法

ELECTRON MICROSCOPY (3.6 Å)