8ZDR
Cryo-EM structure of the Cas9d-sgRNA-target DNA complex
Summary for 8ZDR
| Entry DOI | 10.2210/pdb8zdr/pdb |
| EMDB information | 60006 |
| Descriptor | RNA (144-MER), DNA (32-MER), a protein, ... (5 entities in total) |
| Functional Keywords | a protein complex, antiviral protein |
| Biological source | metagenome More |
| Total number of polymer chains | 4 |
| Total formula weight | 160551.94 |
| Authors | |
| Primary citation | Yang, J.,Wang, T.,Huang, Y.,Long, Z.,Li, X.,Zhang, S.,Zhang, L.,Liu, Z.,Zhang, Q.,Sun, H.,Zhang, M.,Yin, H.,Liu, Z.,Zhang, H. Insights into the compact CRISPR-Cas9d system. Nat Commun, 16:2462-2462, 2025 Cited by PubMed Abstract: Cas9d, the smallest known member of the Cas9 family, employs a compact domain architecture for effective target cleavage. However, the underlying mechanism remains unclear. Here, we present the cryo-EM structures of the Cas9d-sgRNA complex in both target-free and target-bound states. Biochemical assays elucidated the PAM recognition and DNA cleavage mechanisms of Cas9d. Structural comparisons revealed that at least 17 base pairs in the guide-target heteroduplex is required for nuclease activity. Beyond its typical role as an adaptor between Cas9 enzymes and targets, the sgRNA also provides structural support and functional regulation for Cas9d. A segment of the sgRNA scaffold interacts with the REC domain to form a functional target recognition module. Upon target binding, this module undergoes a coordinated conformational rearrangement, enabling heteroduplex propagation and facilitating nuclease activity. This hybrid functional module precisely monitors heteroduplex complementarity, resulting in a lower mismatch tolerance compared to SpyCas9. Moreover, structure-guided engineering in both the sgRNA and Cas9d protein led to a more compact Cas9 system with well-maintained nuclease activity. Altogether, our findings provide insights into the target recognition and cleavage mechanisms of Cas9d and shed light on the development of high-fidelity mini-CRISPR tools. PubMed: 40075056DOI: 10.1038/s41467-025-57455-9 PDB entries with the same primary citation |
| Experimental method | ELECTRON MICROSCOPY (2.65 Å) |
Structure validation
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