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Yorodumi- EMDB-1710: Cryo-EM 3D model of the icosahedral particle composed of Rous sar... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-1710 | |||||||||
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Title | Cryo-EM 3D model of the icosahedral particle composed of Rous sarcoma virus capsid protein pentamers | |||||||||
Map data | This is a 3D reconstruction of the icosahedral particle composed of Rous sarcoma virus capsid protein pentamers | |||||||||
Sample |
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Function / homology | Function and homology information host cell nucleoplasm / viral procapsid maturation / host cell nucleolus / Hydrolases; Acting on peptide bonds (peptidases); Aspartic endopeptidases / viral capsid / aspartic-type endopeptidase activity / nucleic acid binding / structural constituent of virion / host cell plasma membrane / proteolysis ...host cell nucleoplasm / viral procapsid maturation / host cell nucleolus / Hydrolases; Acting on peptide bonds (peptidases); Aspartic endopeptidases / viral capsid / aspartic-type endopeptidase activity / nucleic acid binding / structural constituent of virion / host cell plasma membrane / proteolysis / zinc ion binding / membrane Similarity search - Function | |||||||||
Biological species | Rous sarcoma virus - Prague C | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 18.3 Å | |||||||||
Authors | Hyun JK / Radjainia M / Kingston RL / Mitra AK | |||||||||
Citation | Journal: J Biol Chem / Year: 2010 Title: Proton-driven assembly of the Rous Sarcoma virus capsid protein results in the formation of icosahedral particles. Authors: Jae-Kyung Hyun / Mazdak Radjainia / Richard L Kingston / Alok K Mitra / Abstract: In a mature and infectious retroviral particle, the capsid protein (CA) forms a shell surrounding the genomic RNA and the replicative machinery of the virus. The irregular nature of this capsid shell ...In a mature and infectious retroviral particle, the capsid protein (CA) forms a shell surrounding the genomic RNA and the replicative machinery of the virus. The irregular nature of this capsid shell precludes direct atomic resolution structural analysis. CA hexamers and pentamers are the fundamental building blocks of the capsid, however the pentameric state, in particular, remains poorly characterized. We have developed an efficient in vitro protocol for studying the assembly of Rous sarcoma virus (RSV) CA that involves mild acidification and produces structures modeling the authentic viral capsid. These structures include regular spherical particles with T = 1 icosahedral symmetry, built from CA pentamers alone. These particles were subject to cryoelectron microscopy (cryo-EM) and image processing, and a pseudo-atomic model of the icosahedron was created by docking atomic structures of the constituent CA domains into the cryo-EM-derived three-dimensional density map. The N-terminal domain (NTD) of CA forms pentameric turrets, which decorate the surface of the icosahedron, while the C-terminal domain (CTD) of CA is positioned underneath, linking the pentamers. Biophysical analysis of the icosahedral particle preparation reveals that CA monomers and icosahedra are the only detectable species and that these exist in reversible equilibrium at pH 5. These same acidic conditions are known to promote formation of a RSV CA CTD dimer, present within the icosahedral particle, which facilitates capsid assembly. The results are consistent with a model in which RSV CA assembly is a nucleation-limited process driven by very weak protein-protein interactions. | |||||||||
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_1710.map.gz | 7.4 MB | EMDB map data format | |
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Header (meta data) | emd-1710-v30.xml emd-1710.xml | 10.7 KB 10.7 KB | Display Display | EMDB header |
Images | map_EMD-1710.tif | 758.4 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-1710 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-1710 | HTTPS FTP |
-Validation report
Summary document | emd_1710_validation.pdf.gz | 236.8 KB | Display | EMDB validaton report |
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Full document | emd_1710_full_validation.pdf.gz | 235.9 KB | Display | |
Data in XML | emd_1710_validation.xml.gz | 5.6 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1710 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-1710 | HTTPS FTP |
-Related structure data
Related structure data | 2x8qMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_1710.map.gz / Format: CCP4 / Size: 15.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | This is a 3D reconstruction of the icosahedral particle composed of Rous sarcoma virus capsid protein pentamers | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.5 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 : Icosahedral particles composed of Rous sarcoma virus capsid protein
Entire | Name: Icosahedral particles composed of Rous sarcoma virus capsid protein |
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Components |
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-Supramolecule #1000: Icosahedral particles composed of Rous sarcoma virus capsid protein
Supramolecule | Name: Icosahedral particles composed of Rous sarcoma virus capsid protein type: sample / ID: 1000 Details: The icosahedral particles were assembled in vitro, by transferring recombinant Rous sarcoma virus capsid protein monomers into high salt, mildly acidic buffer Oligomeric state: Icosahedral particle containing 12 CA pentamers (i.e. 60 monomers) Number unique components: 1 |
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Molecular weight | Theoretical: 1.5 MDa |
-Macromolecule #1: Capsid protein p27
Macromolecule | Name: Capsid protein p27 / type: protein_or_peptide / ID: 1 / Name.synonym: Capsid protein p27 / Number of copies: 60 Oligomeric state: Icosahedral particle composed of 12 protein pentamers Recombinant expression: Yes |
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Source (natural) | Organism: Rous sarcoma virus - Prague C |
Molecular weight | Theoretical: 1.53 MDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.2 mg/mL |
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Buffer | pH: 5 Details: 0.1M citric acid, 5mM MOPS/KOH, 725mM NaCl, 0.25mM Na azide, 0.125mM TCEP-HCl |
Grid | Details: Holey carbon 400 mesh copper grid |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 85 K / Instrument: FEI VITROBOT MARK IV / Details: Vitrification instrument: Vitrobot Mark IV / Method: Blot for 5 seconds before plunging |
-Electron microscopy
Microscope | FEI TECNAI 12 |
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Temperature | Average: 103 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 60,000 - 140,000 times magnification using live fft |
Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: NIKON SUPER COOLSCAN 9000 / Digitization - Sampling interval: 10.5 µm / Number real images: 21 / Average electron dose: 18 e/Å2 / Bits/pixel: 8 |
Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 42000 |
Sample stage | Specimen holder: Side entry liquid nitrogen-cooled cryo specimen holder Specimen holder model: GATAN LIQUID NITROGEN |
-Image processing
CTF correction | Details: Each micrograph |
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Final reconstruction | Applied symmetry - Point group: I (icosahedral) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 18.3 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: Bsoft, PFT2, EM3DR2 Details: The digitized micrographs were processed using Bsoft. Orientation and origin search of the particles and 3D reconstruction were performed using PFT2 and EM3DR, respectively Number images used: 1310 |
-Atomic model buiding 1
Initial model | (PDB ID: , ) |
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Software | Name: Chimera,Sculptor |
Details | Protocol: Rigid body. The domain structures were manually fitted into the 3D reconstruction, and then the fitting was refined using Sculptor |
Refinement | Space: RECIPROCAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation |
Output model | PDB-2x8q: |