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- PDB-3j9s: Single particle cryo-EM structure of rotavirus VP6 at 2.6 Angstro... -
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Basic information
Entry | Database: PDB / ID: 3j9s | ||||||
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Title | Single particle cryo-EM structure of rotavirus VP6 at 2.6 Angstrom resolution | ||||||
![]() | Intermediate capsid protein VP6 | ||||||
![]() | VIRAL PROTEIN / rotavirus / virus | ||||||
Function / homology | ![]() viral intermediate capsid / T=13 icosahedral viral capsid / host cell surface receptor binding / fusion of virus membrane with host plasma membrane / viral envelope / structural molecule activity / metal ion binding Similarity search - Function | ||||||
Biological species | ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.6 Å | ||||||
![]() | Grant, T. / Grigorieff, N. | ||||||
![]() | ![]() Title: Measuring the optimal exposure for single particle cryo-EM using a 2.6 Å reconstruction of rotavirus VP6. Authors: Timothy Grant / Nikolaus Grigorieff / ![]() Abstract: Biological specimens suffer radiation damage when imaged in an electron microscope, ultimately limiting the attainable resolution. At a given resolution, an optimal exposure can be defined that ...Biological specimens suffer radiation damage when imaged in an electron microscope, ultimately limiting the attainable resolution. At a given resolution, an optimal exposure can be defined that maximizes the signal-to-noise ratio in the image. Using a 2.6 Å resolution single particle cryo-EM reconstruction of rotavirus VP6, determined from movies recorded with a total exposure of 100 electrons/Å(2), we obtained accurate measurements of optimal exposure values over a wide range of resolutions. At low and intermediate resolutions, our measured values are considerably higher than obtained previously for crystalline specimens, indicating that both images and movies should be collected with higher exposures than are generally used. We demonstrate a method of using our optimal exposure values to filter movie frames, yielding images with improved contrast that lead to higher resolution reconstructions. This 'high-exposure' technique should benefit cryo-EM work on all types of samples, especially those of relatively low-molecular mass. | ||||||
History |
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Structure visualization
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 79.2 KB | Display | ![]() |
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PDB format | ![]() | 60 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 6272MC ![]() 6464C 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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Symmetry | Point symmetry: (Schoenflies symbol: C3 (3 fold cyclic)) |
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Components
#1: Protein | Mass: 44905.676 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() | ||
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#2: Chemical | ChemComp-ZN / | ||
#3: Chemical | ChemComp-CL / | ||
#4: Chemical | #5: Water | ChemComp-HOH / | |
-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: Bovine rotavirus VP6 / Type: VIRUS |
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Specimen | Conc.: 2.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: C-Flat 1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK II / Cryogen name: ETHANE / Temp: 120 K / Humidity: 80 % Details: Blot for 4-6 seconds before plunging into liquid ethane (FEI VITROBOT MARK II). Method: Blot for 4-6 seconds |
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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 / Date: Aug 13, 2014 |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 29000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 400 nm / Cs: 2.7 mm |
Specimen holder | Specimen holder type: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 100 e/Å2 / Film or detector model: GATAN K2 (4k x 4k) Details: 130 frames, 0.1 seconds per frame, 100 e/A2, super resolution |
Radiation wavelength | Relative weight: 1 |
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Processing
EM software |
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CTF correction | Details: each particle | ||||||||||||||||||
Symmetry | Point symmetry: I (icosahedral) | ||||||||||||||||||
3D reconstruction | Method: projection matching / Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 4000 / Nominal pixel size: 1.023 Å / Actual pixel size: 1.023 Å Details: Final map is a 13-fold average of VP6 trimers from the asymmetric unit of the reconstruction of the whole capsid. Data at resolutions higher than 15A were not used for alignments. Symmetry type: POINT | ||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL Details: REFINEMENT PROTOCOL--flexible DETAILS--Model was refined by real space refinement using COOT and all restraints. | ||||||||||||||||||
Atomic model building | PDB-ID: 1QHD Pdb chain-ID: A | ||||||||||||||||||
Refinement step | Cycle: LAST
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