論文：Structure of the miniature type V-F CRISPR-Cas effector enzyme
- クライオ電⼦顕微鏡を⽤いた単粒⼦解析法により、CRISPR-Casヌクレアーゼの一つであるtype V-F Cas12fとガイドRNA、標的DNAの複合体の構造を明らかにした。
- CRISPR-Casヌクレアーゼは複数知られているが、type V-F Cas12fは最も小さくてコンパクトであるにもかかわらず十分な活性をもっている。このタンパク質はホモ２量体であるにもかかわらず、コンフォメーションを変えることで異なる役割を分担する、“asymmetric homodimer”として機能することを明らかにした。
Abstract
RNA-guided DNA endonucleases derived from CRISPR-Cas adaptive immune systems are widely used as powerful genome-engineering tools. Among the diverse CRISPR-Cas nucleases, the type V-F Cas12f (also known as Cas14) proteins are exceptionally compact and associate with a guide RNA to cleave single- and double-stranded DNA targets. Here, we report the cryo-electron microscopy structure of Cas12f1 (also known as Cas14a) in complex with a guide RNA and its target DNA. Unexpectedly, the structure revealed that two Cas12f1 molecules assemble with the single guide RNA to recognize the double-stranded DNA target. Each Cas12f1 protomer adopts a different conformation and plays distinct roles in nucleic acid recognition and DNA cleavage, thereby explaining how the miniature Cas12f1 enzyme achieves RNA-guided DNA cleavage as an ‘‘asymmetric homodimer.’’ Our findings augment the mechanistic understanding of diverse CRISPR-Cas nucleases and provide a framework for the development of compact genome-engineering tools critical for therapeutic genome editing.
分解能
3.3 Å
Citation
- Takeda, R. Nakagawa, S. Okazaki, H. Hirano, K. Kobayashi, T. Kusakizako, T. Nishizawa, K. Yamashita, H. Nishimasu, O. Nureki. Mol. Cell. Feb 4;81(3):558-570 (2021).
