9VJ8

Type II-A CRISPR integrase complex, apo form

Summary for 9VJ8

• Entry DOI: 10.2210/pdb9vj8/pdb
• EMDB information: 65106
• Descriptor: Type II-A CRISPR-associated protein Csn2, CRISPR-associated endonuclease Cas1, CRISPR-associated endoribonuclease Cas2, ... (5 entities in total)
• Functional Keywords: crispr, cas9, cas1, cas2, csn2, dna binding protein
• Biological source: Enterococcus faecalis; More
• Total number of polymer chains: 22
• Total formula weight: 834925.44

Authors

Li, Z.,Li, Y.,Wu, Q.,Lu, M.,Xiao, Y. (deposition date: 2025-06-19, release date: 2026-04-01)

Primary citation

Li, Z.,Li, Y.,Kong, J.,Wu, Q.,Huang, P.,Zhang, Y.,Wu, W.,Chen, M.,Liu, Y.,Lin, H.,Hou, L.,Liu, G.,Zeng, T.,He, Y.,Hu, C.,Yang, Z.,Lu, M.,Luo, M.,Xiao, Y.
Structural basis for Cas9-directed spacer acquisition in type II-A CRISPR-Cas systems.
Mol.Cell, 86:805-816.e4, 2026

PubMed Abstract: CRISPR-Cas systems confer prokaryotic immunity by integrating foreign DNA (prespacers) into host arrays. Type II-A systems employ Cas9 for protospacer-adjacent motif (PAM) recognition and coordinate with Csn2 and the Cas1-Cas2 integrase during spacer acquisition, yet their structural basis remains unresolved. Here, we report cryo-electron microscopy (cryo-EM) structures of the Enterococcus faecalis Cas9-Csn2-Cas1-Cas2 supercomplex in apo and DNA-bound states. The apo state (Cas9₂-Csn2₈-Cas1₈-Cas2₄) is a resting complex, while DNA binding forms a prespacer-catching complex threading DNA through Csn2's channel, enabling Cas9 to interrogate the PAM sequence while sliding along the DNA. Cas9 and Csn2 jointly define a 30-bp DNA segment matching the prespacer length. Cas9 dissociation triggers structural reconfiguration of the Csn2-Cas1-Cas2 assembly. This exposes the PAM-proximal DNA, allowing Cas1-Cas2 to bind the exposed site for subsequent prespacer processing and directional integration. These findings reveal how Cas9, Csn2, and Cas1-Cas2 couple PAM recognition with prespacer selection, ensuring fidelity during adaptation.

PubMed: 41702404
DOI: 10.1016/j.molcel.2026.01.024

Experimental method

ELECTRON MICROSCOPY (3.9 Å)