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TitleStructure of the miniature type V-F CRISPR-Cas effector enzyme.
Journal, issue, pagesMol Cell, Vol. 81, Issue 3, Page 558-570.e3, Year 2021
Publish dateFeb 4, 2021
AuthorsSatoru N Takeda / Ryoya Nakagawa / Sae Okazaki / Hisato Hirano / Kan Kobayashi / Tsukasa Kusakizako / Tomohiro Nishizawa / Keitaro Yamashita / Hiroshi Nishimasu / Osamu Nureki /
PubMed AbstractRNA-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 ...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.
External linksMol Cell / PubMed:33333018
MethodsEM (single particle)
Resolution3.3 Å
Structure data

30299

7c7l

EMDB-30299, PDB-7c7l:
Cryo-EM structure of the Cas12f1-sgRNA-target DNA complex
Method: EM (single particle) / Resolution: 3.3 Å

Chemicals

ChemComp-ZN:
Unknown entry

Source
  • uncultured archaeon (environmental samples)
  • synthetic construct (others)
KeywordsRNA BINDING PROTEIN/RNA/DNA / Cas12f / Cas14 / sgRNA / target DNA / CRISPR / RNA BINDING PROTEIN-RNA-DNA complex

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