9H21

Cas9 in complex with tracrRNA and crRNA, Streptococcus thermophilus DGCC 7710 CRISPR3 system

Summary for 9H21

• Entry DOI: 10.2210/pdb9h21/pdb
• EMDB information: 51787
• Descriptor: CRISPR-associated endonuclease Cas9, crRNA (5'-R(P*GP*AP*AP*GP*AP*GP*GP*AP*CP*AP*GP*UP*UP*UP*UP*AP*GP*AP*GP*CP*UP*G)-3'), tracrRNA (62-MER), ... (4 entities in total)
• Functional Keywords: cas9, crispr-cas, pam, spacer acquisition, rna binding protein
• Biological source: Streptococcus thermophilus DGCC 7710; More
• Total number of polymer chains: 3
• Total formula weight: 200267.05

Authors

Sasnauskas, G.,Gaizauskaite, U.,Tamulaitiene, G. (deposition date: 2024-10-10, release date: 2026-02-18, Last modification date: 2026-03-18)

Primary citation

Gaizauskaite, U.,Tamulaitiene, G.,Silanskas, A.,Gasiunas, G.,Siksnys, V.,Sasnauskas, G.
Structural insights into Cas9-mediated prespacer selection in CRISPR-Cas adaptation.
Mol.Cell, 86:791-804.e9, 2026

PubMed Abstract: During CRISPR-Cas adaptation, prokaryotic cells become immunized by the insertion of foreign DNA fragments, termed spacers, into the host genome to serve as templates for RNA-guided immunity. Spacer acquisition relies on the Cas1-Cas2 integrase and accessory proteins, which select DNA sequences flanked by the protospacer adjacent motif (PAM) and insert them into the CRISPR array. It has been shown that in type II-A systems, selection of PAM-proximal prespacers is mediated by the effector nuclease Cas9, which forms a "supercomplex" with the Cas1-Cas2 integrase and the Csn2 protein. Here, we present cryo-electron microscopy structures of the Streptococcus thermophilus type II-A prespacer selection supercomplex in the DNA-scanning and two distinct PAM-bound configurations, providing insights into the mechanism of Cas9-mediated prespacer selection in type II-A CRISPR-Cas systems. Repurposing Cas9 by the CRISPR adaptation machinery for prespacer selection, as characterized here, demonstrates Cas9 plasticity and expands our knowledge of Cas9 biology.

PubMed: 41702403
DOI: 10.1016/j.molcel.2026.01.022

Experimental method

ELECTRON MICROSCOPY (3.33 Å)