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- PDB-6o7i: Cryo-EM structure of Csm-crRNA-target RNA ternary bigger complex ... -
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Basic information
Entry | Database: PDB / ID: 6o7i | ||||||
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Title | Cryo-EM structure of Csm-crRNA-target RNA ternary bigger complex in complex with cA4 in type III-A CRISPR-Cas system | ||||||
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![]() | immune system/rna / cryo-EM structure / Csm-crRNA-target RNA ternary bigger complex in complex with CA4 / Type III CRISPR-Cas systerm / IMMUNE SYSTEM / immune system-rna-dna complex / immune system-rna complex | ||||||
Function / homology | ![]() exonuclease activity / Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases / transferase activity / endonuclease activity / defense response to virus / Hydrolases; Acting on ester bonds / RNA binding / ATP binding / identical protein binding Similarity search - Function | ||||||
Biological species | ![]() ![]() synthetic construct (others) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å | ||||||
![]() | Jia, N. / Patel, D.J. | ||||||
![]() | ![]() Title: Second Messenger cA Formation within the Composite Csm1 Palm Pocket of Type III-A CRISPR-Cas Csm Complex and Its Release Path. Authors: Ning Jia / Roger Jones / George Sukenick / Dinshaw J Patel / ![]() Abstract: Target RNA binding to crRNA-bound type III-A CRISPR-Cas multi-subunit Csm surveillance complexes activates cyclic-oligoadenylate (cA) formation from ATP subunits positioned within the composite pair ...Target RNA binding to crRNA-bound type III-A CRISPR-Cas multi-subunit Csm surveillance complexes activates cyclic-oligoadenylate (cA) formation from ATP subunits positioned within the composite pair of Palm domain pockets of the Csm1 subunit. The generated cA second messenger in turn targets the CARF domain of trans-acting RNase Csm6, triggering its HEPN domain-based RNase activity. We have undertaken cryo-EM studies on multi-subunit Thermococcus onnurineus Csm effector ternary complexes, as well as X-ray studies on Csm1-Csm4 cassette, both bound to substrate (AMPPNP), intermediates (pppA), and products (cA), to decipher mechanistic aspects of cA formation and release. A network of intermolecular hydrogen bond alignments accounts for the observed adenosine specificity, with ligand positioning dictating formation of linear pppA intermediates and subsequent cA formation by cyclization. We combine our structural results with published functional studies to highlight mechanistic insights into the role of the Csm effector complex in mediating the cA signaling pathway. | ||||||
History |
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 462.8 KB | Display | ![]() |
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PDB format | ![]() | 366.3 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Summary document | ![]() | 826.2 KB | Display | ![]() |
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Full document | ![]() | 853.6 KB | Display | |
Data in XML | ![]() | 69.8 KB | Display | |
Data in CIF | ![]() | 111.9 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 0642MC ![]() 0640C ![]() 0641C ![]() 6o73C ![]() 6o74C ![]() 6o75C ![]() 6o78C ![]() 6o79C ![]() 6o7bC ![]() 6o7dC ![]() 6o7eC ![]() 6o7hC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Components
-Protein , 5 types, 8 molecules ABJCDKEF
#1: Protein | Mass: 89706.594 Da / Num. of mol.: 1 / Mutation: D589A Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: NA1 / Gene: csm1, cas10, TON_0893 / Production host: ![]() ![]() References: UniProt: B6YWB8, Hydrolases; Acting on ester bonds, Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases | ||||||
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#2: Protein | Mass: 21210.293 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: NA1 / Gene: TON_0894 / Production host: ![]() ![]() #3: Protein | Mass: 32809.012 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: NA1 / Production host: ![]() ![]() #4: Protein | | Mass: 32345.061 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: NA1 / Gene: TON_0896 / Production host: ![]() ![]() #8: Protein | | Mass: 40588.410 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: NA1 / Gene: TON_0897 / Production host: ![]() ![]() |
-RNA chain , 3 types, 3 molecules GHI
#5: RNA chain | Mass: 12463.438 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
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#6: RNA chain | Mass: 12750.658 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) ![]() ![]() |
#7: RNA chain | Mass: 1271.866 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
-Non-polymers , 1 types, 4 molecules ![](data/chem/img/ZN.gif)
#9: Chemical | ChemComp-ZN / |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Csm-crRNA-target RNA complex / Type: COMPLEX / Entity ID: #1-#7 / Source: RECOMBINANT |
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Molecular weight | Value: 0.3 MDa / Experimental value: YES |
Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 8.8 / Details: 20 mM Tris-HCl, pH 8.8, 250 mM NaCl, 2 mM DTT |
Buffer component | Formula: Tris |
Specimen | Conc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % |
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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 |
Image recording | Electron dose: 1.35 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
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Processing
EM software | Name: RELION / Version: 2.1 / Category: 3D reconstruction |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION |
3D reconstruction | Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 41472 / Symmetry type: POINT |