CRISPR-associated protein, MJ0385 / CRISPR-associated protein (Cas_Csa4) / : / CRISPR-associated protein, Cas5a type / : / CRISPR-associated protein Cas7/Cst2/DevR, subtype I-a/Apern / CRISPR-associated protein Cas7/Cst2/DevR / CRISPR-associated negative auto-regulator DevR/Csa2 / CRISPR-associated protein Cas5, N-terminal 類似検索 - ドメイン・相同性
Type I-A CRISPR-associated protein Cas5 / Uncharacterized protein / Type I-A CRISPR-associated protein Cas7/Csa2 / Type I-A CRISPR-associated protein Cas8a2/Csa4 類似検索 - 構成要素
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
GM118117
米国
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
GM118160
米国
Swiss National Science Foundation
NCCR
スイス
引用
ジャーナル: Mol Cell / 年: 2022 タイトル: Allosteric control of type I-A CRISPR-Cas3 complexes and establishment as effective nucleic acid detection and human genome editing tools. 著者: Chunyi Hu / Dongchun Ni / Ki Hyun Nam / Sonali Majumdar / Justin McLean / Henning Stahlberg / Michael P Terns / Ailong Ke / 要旨: Type I CRISPR-Cas systems typically rely on a two-step process to degrade DNA. First, an RNA-guided complex named Cascade identifies the complementary DNA target. The helicase-nuclease fusion enzyme ...Type I CRISPR-Cas systems typically rely on a two-step process to degrade DNA. First, an RNA-guided complex named Cascade identifies the complementary DNA target. The helicase-nuclease fusion enzyme Cas3 is then recruited in trans for processive DNA degradation. Contrary to this model, here, we show that type I-A Cascade and Cas3 function as an integral effector complex. We provide four cryoelectron microscopy (cryo-EM) snapshots of the Pyrococcus furiosus (Pfu) type I-A effector complex in different stages of DNA recognition and degradation. The HD nuclease of Cas3 is autoinhibited inside the effector complex. It is only allosterically activated upon full R-loop formation, when the entire targeted region has been validated by the RNA guide. The mechanistic insights inspired us to convert Pfu Cascade-Cas3 into a high-sensitivity, low-background, and temperature-activated nucleic acid detection tool. Moreover, Pfu CRISPR-Cas3 shows robust bi-directional deletion-editing activity in human cells, which could find usage in allele-specific inactivation of disease-causing mutations.