8BD5

Cas12k-sgRNA-dsDNA-S15-TniQ-TnsC transposon recruitment complex

Summary for 8BD5

• Entry DOI: 10.2210/pdb8bd5/pdb
• EMDB information: 15975
• Descriptor: ShCas12k, ZINC ION, sgRNA, ... (10 entities in total)
• Functional Keywords: cas12k, sgrna, s15, tniq, tnsc, crispr-cas, tn7-like transposons, transposition, rna binding protein
• Biological source: Scytonema hofmannii; More
• Total number of polymer chains: 13
• Total formula weight: 444841.26

Authors

Schmitz, M.,Querques, I.,Oberli, S.,Chanez, C.,Jinek, M. (deposition date: 2022-10-18, release date: 2022-12-28, Last modification date: 2024-07-24)

Primary citation

Schmitz, M.,Querques, I.,Oberli, S.,Chanez, C.,Jinek, M.
Structural basis for the assembly of the type V CRISPR-associated transposon complex.
Cell, 185:4999-, 2022

PubMed Abstract: CRISPR-Cas systems have been co-opted by Tn7-like transposable elements to direct RNA-guided transposition. Type V-K CRISPR-associated transposons rely on the concerted activities of the pseudonuclease Cas12k, the AAA+ ATPase TnsC, the Zn-finger protein TniQ, and the transposase TnsB. Here we present a cryo-electron microscopic structure of a target DNA-bound Cas12k-transposon recruitment complex comprised of RNA-guided Cas12k, TniQ, a polymeric TnsC filament and, unexpectedly, the ribosomal protein S15. Complex assembly, mediated by a network of interactions involving the guide RNA, TniQ, and S15, results in R-loop completion. TniQ contacts two TnsC protomers at the Cas12k-proximal filament end, likely nucleating its polymerization. Transposition activity assays corroborate our structural findings, implying that S15 is a bona fide component of the type V crRNA-guided transposon machinery. Altogether, our work uncovers key mechanistic aspects underpinning RNA-mediated assembly of CRISPR-associated transposons to guide their development as programmable tools for site-specific insertion of large DNA payloads.

PubMed: 36435179
DOI: 10.1016/j.cell.2022.11.009

Experimental method

ELECTRON MICROSCOPY (3.3 Å)