7M5O

Cryo-EM structure of CasPhi-2 (Cas12j) bound to crRNA

7M5O の概要

• エントリーDOI: 10.2210/pdb7m5o/pdb
• EMDBエントリー: 23678
• 分子名称: CasPhi, crRNA, ZINC ION (3 entities in total)
• 機能のキーワード: crispr, casphi, cas12j, nuclease, crrna, rnp, complex, viral protein-rna complex, viral protein/rna
• 由来する生物種: Biggievirus Mos11 (Biggievirus); 詳細
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 100674.25

構造登録者

Pausch, P.,Soczek, K.,Nogales, E.,Doudna, J. (登録日: 2021-03-24, 公開日: 2021-08-04, 最終更新日: 2024-05-29)

主引用文献

Pausch, P.,Soczek, K.M.,Herbst, D.A.,Tsuchida, C.A.,Al-Shayeb, B.,Banfield, J.F.,Nogales, E.,Doudna, J.A.
DNA interference states of the hypercompact CRISPR-Cas Phi effector.
Nat.Struct.Mol.Biol., 28:652-661, 2021

PubMed Abstract: CRISPR-CasΦ, a small RNA-guided enzyme found uniquely in bacteriophages, achieves programmable DNA cutting as well as genome editing. To investigate how the hypercompact enzyme recognizes and cleaves double-stranded DNA, we determined cryo-EM structures of CasΦ (Cas12j) in pre- and post-DNA-binding states. The structures reveal a streamlined protein architecture that tightly encircles the CRISPR RNA and DNA target to capture, unwind and cleave DNA. Comparison of the pre- and post-DNA-binding states reveals how the protein rearranges for DNA cleavage upon target recognition. On the basis of these structures, we created and tested mutant forms of CasΦ that cut DNA up to 20-fold faster relative to wild type, showing how this system may be naturally attenuated to improve the fidelity of DNA interference. The structural and mechanistic insights into how CasΦ binds and cleaves DNA should allow for protein engineering for both in vitro diagnostics and genome editing.

PubMed: 34381246
DOI: 10.1038/s41594-021-00632-3

実験手法

ELECTRON MICROSCOPY (3.54 Å)