Molecular basis and engineering of miniature Cas12f with C-rich PAM specificity.
Nat Chem Biol
; 20(2): 180-189, 2024 Feb.
Article
en En
| MEDLINE
| ID: mdl-37697004
ABSTRACT
CRISPR-Cas12f nucleases are currently one of the smallest genome editors, exhibiting advantages for efficient delivery via cargo-size-limited adeno-associated virus delivery vehicles. Most characterized Cas12f nucleases recognize similar T-rich protospacer adjacent motifs (PAMs) for DNA targeting, substantially restricting their targeting scope. Here we report the cryogenic electron microscopy structure and engineering of a miniature Clostridium novyi Cas12f1 nuclease (CnCas12f1, 497 amino acids) with rare C-rich PAM specificity. Structural characterizations revealed detailed PAM recognition, asymmetric homodimer formation and single guide RNA (sgRNA) association mechanisms. sgRNA engineering transformed CRISPR-CnCas12f1, which initially was incapable of genome targeting in bacteria, into an effective genome editor in human cells. Our results facilitate further understanding of CRISPR-Cas12f1 working mechanism and expand the mini-CRISPR toolbox.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Sistemas CRISPR-Cas
/
ARN Guía de Sistemas CRISPR-Cas
Límite:
Humans
Idioma:
En
Revista:
Nat Chem Biol
Asunto de la revista:
BIOLOGIA
/
QUIMICA
Año:
2024
Tipo del documento:
Article
País de afiliación:
China