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Open data
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Basic information
Entry | Database: PDB / ID: 8wmc | |||||||||
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Title | Cryo-EM structure of DiCas7-11-crRNA in complex with regulator | |||||||||
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![]() | IMMUNE SYSTEM / CRISPR-Cas / Gene editing / TPR-CHAT | |||||||||
Function / homology | CHAT domain / CHAT domain / CRISPR type III-associated protein / RAMP superfamily / defense response to virus / RNA / RNA (> 10) / CRISPR-associated RAMP family protein / CHAT domain-containing protein![]() | |||||||||
Biological species | ![]() ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å | |||||||||
![]() | Ma, H.Y. / Tang, X.D. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural basis of negative regulation of CRISPR-Cas7-11 by TPR-CHAT. Authors: Tian Hong / Qinghua Luo / Haiyun Ma / Xin Wang / Xinqiong Li / Chongrong Shen / Jie Pang / Yan Wang / Yuejia Chen / Changbin Zhang / Zhaoming Su / Haohao Dong / Xiaodi Tang / ![]() Abstract: CRISPR‒Cas7-11 is a Type III-E CRISPR-associated nuclease that functions as a potent RNA editing tool. Tetratrico-peptide repeat fused with Cas/HEF1-associated signal transducer (TPR-CHAT) acts as ...CRISPR‒Cas7-11 is a Type III-E CRISPR-associated nuclease that functions as a potent RNA editing tool. Tetratrico-peptide repeat fused with Cas/HEF1-associated signal transducer (TPR-CHAT) acts as a regulatory protein that interacts with CRISPR RNA (crRNA)-bound Cas7-11 to form a CRISPR-guided caspase complex (Craspase). However, the precise modulation of Cas7-11's nuclease activity by TPR-CHAT to enhance its utility requires further study. Here, we report cryo-electron microscopy (cryo-EM) structures of Desulfonema ishimotonii (Di) Cas7-11-crRNA, complexed with or without the full length or the N-terminus of TPR-CHAT. These structures unveil the molecular features of the Craspase complex. Structural analysis, combined with in vitro nuclease assay and electrophoretic mobility shift assay, reveals that DiTPR-CHAT negatively regulates the activity of DiCas7-11 by preventing target RNA from binding through the N-terminal 65 amino acids of DiTPR-CHAT (DiTPR-CHAT). Our work demonstrates that DiTPR-CHAT can function as a small unit of DiCas7-11 regulator, potentially enabling safe applications to prevent overcutting and off-target effects of the CRISPR‒Cas7-11 system. | |||||||||
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 390.9 KB | Display | ![]() |
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PDB format | ![]() | 306.8 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.6 MB | Display | ![]() |
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Full document | ![]() | 1.6 MB | Display | |
Data in XML | ![]() | 64.1 KB | Display | |
Data in CIF | ![]() | 96.6 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 37649MC ![]() 8wm4C ![]() 8wmiC ![]() 8wmlC C: citing same article ( M: map data used to model this data |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Components
#1: Protein | Mass: 88311.875 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() | ||
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#2: Protein | Mass: 183933.000 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() | ||
#3: RNA chain | Mass: 12134.133 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() | ||
#4: Chemical | ChemComp-ZN / Has ligand of interest | Y | |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: CELL / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Ternary complex of Cas7-11_crRNA_TPR-CHAT / Type: COMPLEX / Entity ID: #2, #1, #3 / Source: RECOMBINANT |
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Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 8 |
Specimen | Conc.: 0.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: NITROGEN |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 3500 nm / Nominal defocus min: 960 nm / Cs: 2.7 mm |
Image recording | Electron dose: 53.7 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) |
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Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
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3D reconstruction | Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 78984 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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