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Yorodumi- EMDB-5719: Electron microscopy of the negatively-stained Cmr complex from Th... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-5719 | |||||||||
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Title | Electron microscopy of the negatively-stained Cmr complex from Thermus thermophilus HB8. | |||||||||
Map data | 3D structure by EM single particle analysis of negatively-stained images | |||||||||
Sample |
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Keywords | adaptive immune system / cmr complex / CRISPR-Cas / Thermus thermophilus | |||||||||
Function / homology | Function and homology information | |||||||||
Biological species | Thermus thermophilus (bacteria) | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 26.0 Å | |||||||||
Authors | Agari Y / Maki-Yonekura S / Staals RH / van Duijin E / Heck AJ / Yonekura K / van der Oost J / Shinkai A | |||||||||
Citation | Journal: Mol Cell / Year: 2013 Title: Structure and activity of the RNA-targeting Type III-B CRISPR-Cas complex of Thermus thermophilus. Authors: Raymond H J Staals / Yoshihiro Agari / Saori Maki-Yonekura / Yifan Zhu / David W Taylor / Esther van Duijn / Arjan Barendregt / Marnix Vlot / Jasper J Koehorst / Keiko Sakamoto / Akiko ...Authors: Raymond H J Staals / Yoshihiro Agari / Saori Maki-Yonekura / Yifan Zhu / David W Taylor / Esther van Duijn / Arjan Barendregt / Marnix Vlot / Jasper J Koehorst / Keiko Sakamoto / Akiko Masuda / Naoshi Dohmae / Peter J Schaap / Jennifer A Doudna / Albert J R Heck / Koji Yonekura / John van der Oost / Akeo Shinkai / Abstract: The CRISPR-Cas system is a prokaryotic host defense system against genetic elements. The Type III-B CRISPR-Cas system of the bacterium Thermus thermophilus, the TtCmr complex, is composed of six ...The CRISPR-Cas system is a prokaryotic host defense system against genetic elements. The Type III-B CRISPR-Cas system of the bacterium Thermus thermophilus, the TtCmr complex, is composed of six different protein subunits (Cmr1-6) and one crRNA with a stoichiometry of Cmr112131445361:crRNA1. The TtCmr complex copurifies with crRNA species of 40 and 46 nt, originating from a distinct subset of CRISPR loci and spacers. The TtCmr complex cleaves the target RNA at multiple sites with 6 nt intervals via a 5' ruler mechanism. Electron microscopy revealed that the structure of TtCmr resembles a "sea worm" and is composed of a Cmr2-3 heterodimer "tail," a helical backbone of Cmr4 subunits capped by Cmr5 subunits, and a curled "head" containing Cmr1 and Cmr6. Despite having a backbone of only four Cmr4 subunits and being both longer and narrower, the overall architecture of TtCmr resembles that of Type I Cascade complexes. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_5719.map.gz | 293.4 KB | EMDB map data format | |
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Header (meta data) | emd-5719-v30.xml emd-5719.xml | 14.5 KB 14.5 KB | Display Display | EMDB header |
Images | emd_5719_1.jpg | 51.1 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-5719 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-5719 | HTTPS FTP |
-Validation report
Summary document | emd_5719_validation.pdf.gz | 78.4 KB | Display | EMDB validaton report |
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Full document | emd_5719_full_validation.pdf.gz | 77.5 KB | Display | |
Data in XML | emd_5719_validation.xml.gz | 494 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5719 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-5719 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_5719.map.gz / Format: CCP4 / Size: 309.6 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | 3D structure by EM single particle analysis of negatively-stained images | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 4.6 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Cmr complex from Thermus thermophilus
Entire | Name: Cmr complex from Thermus thermophilus |
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Components |
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-Supramolecule #1000: Cmr complex from Thermus thermophilus
Supramolecule | Name: Cmr complex from Thermus thermophilus / type: sample / ID: 1000 / Details: sample was monodisperse / Oligomeric state: hetero multimer / Number unique components: 7 |
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Molecular weight | Experimental: 400 KDa / Theoretical: 360 KDa / Method: mass spectorometry |
-Macromolecule #1: Cmr2
Macromolecule | Name: Cmr2 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Thermus thermophilus (bacteria) / Strain: HB8 / Location in cell: cytoplasm |
Molecular weight | Experimental: 65 KDa / Theoretical: 65 KDa |
Sequence | UniProtKB: GGDEF domain-containing protein |
-Macromolecule #2: Cmr3
Macromolecule | Name: Cmr3 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Thermus thermophilus (bacteria) / Strain: HB8 |
Molecular weight | Experimental: 40 KDa / Theoretical: 40 KDa |
Sequence | UniProtKB: CRISPR-associated protein Cmr3 |
-Macromolecule #3: Cmr1
Macromolecule | Name: Cmr1 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Thermus thermophilus (bacteria) / Strain: HB8 |
Molecular weight | Experimental: 40 KDa / Theoretical: 40 KDa |
Sequence | UniProtKB: Type III-B CRISPR module RAMP protein Cmr1 |
-Macromolecule #4: Cmr4
Macromolecule | Name: Cmr4 / type: protein_or_peptide / ID: 4 / Number of copies: 4 / Oligomeric state: tetramer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Thermus thermophilus (bacteria) / Strain: HB8 |
Molecular weight | Experimental: 30 KDa / Theoretical: 30 KDa |
Sequence | UniProtKB: Type III-B CRISPR module RAMP protein Cmr4 |
-Macromolecule #5: Cmr5
Macromolecule | Name: Cmr5 / type: protein_or_peptide / ID: 5 / Number of copies: 3 / Oligomeric state: trimer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Thermus thermophilus (bacteria) / Strain: HB8 |
Molecular weight | Experimental: 10 KDa / Theoretical: 10 KDa |
Sequence | UniProtKB: CRISPR system Cmr subunit Cmr5 |
-Macromolecule #6: Cmr6
Macromolecule | Name: Cmr6 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: Thermus thermophilus (bacteria) / Strain: HB8 |
Molecular weight | Experimental: 40 KDa / Theoretical: 40 KDa |
Sequence | UniProtKB: Type III-B CRISPR module RAMP protein Cmr6 |
-Macromolecule #7: crRNA
Macromolecule | Name: crRNA / type: rna / ID: 7 / Classification: OTHER / Structure: SINGLE STRANDED / Synthetic?: No |
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Source (natural) | Organism: Thermus thermophilus (bacteria) / Strain: HB8 |
Molecular weight | Theoretical: 20 KDa |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.1 mg/mL |
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Buffer | pH: 8 / Details: 20mM Tris-HCl, 150mM NaCl |
Staining | Type: NEGATIVE Details: Grids with adsorbed protein floated on 2% w/v uranyl acetate for 30 seconds. |
Grid | Details: 200 mesh gold grid with thin carbon support |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
-Electron microscopy
Microscope | JEOL 2100 |
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Date | Mar 11, 2011 |
Image recording | Number real images: 105 |
Electron beam | Acceleration voltage: 200 kV / Electron source: LAB6 |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Sample stage | Specimen holder model: JEOL |
-Image processing
Details | The particles were selected interactively at the computer terminal. |
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CTF correction | Details: whole micrograph |
Final reconstruction | Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 26.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: EMAN / Number images used: 8976 |