7C7L

Cryo-EM structure of the Cas12f1-sgRNA-target DNA complex

Summary for 7C7L

• Entry DOI: 10.2210/pdb7c7l/pdb
• EMDB information: 30299
• Descriptor: CRISPR-associated protein Cas14a.1, sgRNA, DNA (40-mer), ... (5 entities in total)
• Functional Keywords: cas12f, cas14, sgrna, target dna, crispr, rna binding protein-rna-dna complex, rna binding protein/rna/dna
• Biological source: uncultured archaeon; More
• Total number of polymer chains: 5
• Total formula weight: 208448.84

Authors

Takeda, N.S.,Nakagawa, R.,Okazaki, S.,Hirano, H.,Kobayashi, K.,Kusakizako, T.,Nishizawa, T.,Yamashita, K.,Nishimasu, H.,Nureki, O. (deposition date: 2020-05-26, release date: 2020-12-23, Last modification date: 2024-03-27)

Primary citation

Takeda, S.N.,Nakagawa, R.,Okazaki, S.,Hirano, H.,Kobayashi, K.,Kusakizako, T.,Nishizawa, T.,Yamashita, K.,Nishimasu, H.,Nureki, O.
Structure of the miniature type V-F CRISPR-Cas effector enzyme.
Mol.Cell, 81:558-570.e3, 2021

PubMed 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.

PubMed: 33333018
DOI: 10.1016/j.molcel.2020.11.035

Experimental method

ELECTRON MICROSCOPY (3.3 Å)