6I1L
Crystal structure of FnCas12a in complex with a crRNA guide and ssDNA target
Summary for 6I1L
| Entry DOI | 10.2210/pdb6i1l/pdb |
| Descriptor | CRISPR-associated endonuclease Cas12a, crRNA (40-MER), ssDNA target strand, ... (7 entities in total) |
| Functional Keywords | crispr-cas12a, fncas12a, crispr-cpf1, fncpf1, hydrolase |
| Biological source | Francisella tularensis subsp. novicida (strain U112) More |
| Total number of polymer chains | 6 |
| Total formula weight | 344862.56 |
| Authors | Jinek, M.,Swarts, D.C. (deposition date: 2018-10-29, release date: 2019-01-23, Last modification date: 2024-10-23) |
| Primary citation | Swarts, D.C.,Jinek, M. Mechanistic Insights into the cis- and trans-Acting DNase Activities of Cas12a. Mol. Cell, 73:589-600.e4, 2019 Cited by PubMed Abstract: CRISPR-Cas12a (Cpf1) is an RNA-guided DNA-cutting nuclease that has been repurposed for genome editing. Upon target DNA binding, Cas12a cleaves both the target DNA in cis and non-target single-stranded DNAs (ssDNAs) in trans. To elucidate the molecular basis for both DNase cleavage modes, we performed structural and biochemical studies on Francisella novicida Cas12a. We show that guide RNA-target strand DNA hybridization conformationally activates Cas12a, triggering its trans-acting, non-specific, single-stranded DNase activity. In turn, cis cleavage of double-stranded DNA targets is a result of protospacer adjacent motif (PAM)-dependent DNA duplex unwinding, electrostatic stabilization of the displaced non-target DNA strand, and ordered sequential cleavage of the non-target and target DNA strands. Cas12a releases the PAM-distal DNA cleavage product and remains bound to the PAM-proximal DNA cleavage product in a catalytically competent, trans-active state. Together, these results provide a revised model for the molecular mechanisms of both the cis- and the trans-acting DNase activities of Cas12a enzymes, enabling their further exploitation as genome editing tools. PubMed: 30639240DOI: 10.1016/j.molcel.2018.11.021 PDB entries with the same primary citation |
| Experimental method | X-RAY DIFFRACTION (2.98 Å) |
Structure validation
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