8AXA

Cryo-EM structure of shCas12k-sgRNA-dsDNA ternary complex (type V-K CRISPR-associated transposon)

8AXA の概要

• エントリーDOI: 10.2210/pdb8axa/pdb
• 関連するPDBエントリー: 8AXB
• EMDBエントリー: 15697 15698 19075 19282 19283 19284 19286
• 分子名称: Cas12k, sgRNA, DNA target strand, ... (5 entities in total)
• 機能のキーワード: crispr, complex, transposition, dna binding protein
• 由来する生物種: Scytonema hofmannii; 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 192631.59

構造登録者

Tenjo-Castano, F.,Sofos, N.,Stella, S.,Fuglsang, A.,Pape, T.,Mesa, P.,Stutzke, L.S.,Temperini, P.,Montoya, G. (登録日: 2022-08-31, 公開日: 2024-04-10, 最終更新日: 2024-07-03)

主引用文献

Tenjo-Castano, F.,Sofos, N.,Stutzke, L.S.,Temperini, P.,Fuglsang, A.,Pape, T.,Mesa, P.,Montoya, G.
Conformational landscape of the type V-K CRISPR-associated transposon integration assembly.
Mol.Cell, 84:2353-2367.e5, 2024

PubMed Abstract: CRISPR-associated transposons (CASTs) are mobile genetic elements that co-opt CRISPR-Cas systems for RNA-guided DNA transposition. CASTs integrate large DNA cargos into the attachment (att) site independently of homology-directed repair and thus hold promise for eukaryotic genome engineering. However, the functional diversity and complexity of CASTs hinder an understanding of their mechanisms. Here, we present the high-resolution cryoelectron microscopy (cryo-EM) structure of the reconstituted ∼1 MDa post-transposition complex of the type V-K CAST, together with different assembly intermediates and diverse TnsC filament lengths, thus enabling the recapitulation of the integration complex formation. The results of mutagenesis experiments probing the roles of specific residues and TnsB-binding sites show that transposition activity can be enhanced and suggest that the distance between the PAM and att sites is determined by the lengths of the TnsB C terminus and the TnsC filament. This singular model of RNA-guided transposition provides a foundation for repurposing the system for genome-editing applications.

PubMed: 38834066
DOI: 10.1016/j.molcel.2024.05.005

実験手法

ELECTRON MICROSCOPY (2.96 Å)