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- EMDB-6404: In situ structures of the segmented genome and RNA polymerase com... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-6404 | |||||||||
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Title | In situ structures of the segmented genome and RNA polymerase complex inside a dsRNA virus | |||||||||
![]() | CPV in the transcribing state | |||||||||
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![]() | cryo-EM / dsRNA genome organization / viral polymerase | |||||||||
Function / homology | ![]() viral genome replication / RNA-dependent RNA polymerase activity / RNA binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.0 Å | |||||||||
![]() | Zhang X / Ding K / Yu XK / Chang W / Sun JC / Zhou ZH | |||||||||
![]() | ![]() Title: In situ structures of the segmented genome and RNA polymerase complex inside a dsRNA virus. Authors: Xing Zhang / Ke Ding / Xuekui Yu / Winston Chang / Jingchen Sun / Z Hong Zhou / ![]() ![]() Abstract: Viruses in the Reoviridae, like the triple-shelled human rotavirus and the single-shelled insect cytoplasmic polyhedrosis virus (CPV), all package a genome of segmented double-stranded RNAs (dsRNAs) ...Viruses in the Reoviridae, like the triple-shelled human rotavirus and the single-shelled insect cytoplasmic polyhedrosis virus (CPV), all package a genome of segmented double-stranded RNAs (dsRNAs) inside the viral capsid and carry out endogenous messenger RNA synthesis through a transcriptional enzyme complex (TEC). By direct electron-counting cryoelectron microscopy and asymmetric reconstruction, we have determined the organization of the dsRNA genome inside quiescent CPV (q-CPV) and the in situ atomic structures of TEC within CPV in both quiescent and transcribing (t-CPV) states. We show that the ten segmented dsRNAs in CPV are organized with ten TECs in a specific, non-symmetric manner, with each dsRNA segment attached directly to a TEC. The TEC consists of two extensively interacting subunits: an RNA-dependent RNA polymerase (RdRP) and an NTPase VP4. We find that the bracelet domain of RdRP undergoes marked conformational change when q-CPV is converted to t-CPV, leading to formation of the RNA template entry channel and access to the polymerase active site. An amino-terminal helix from each of two subunits of the capsid shell protein (CSP) interacts with VP4 and RdRP. These findings establish the link between sensing of environmental cues by the external proteins and activation of endogenous RNA transcription by the TEC inside the virus. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 863.4 KB | ![]() | |
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Header (meta data) | ![]() ![]() | 9.9 KB 9.9 KB | Display Display | ![]() |
Images | ![]() | 541 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 3jb7MC ![]() 6405C ![]() 6406C ![]() 6407C ![]() 6408C ![]() 6409C ![]() 3jb6C M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | CPV in the transcribing state | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.36 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : CPV VP4 + polymerase complex (TEC)
Entire | Name: CPV VP4 + polymerase complex (TEC) |
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Components |
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-Supramolecule #1000: CPV VP4 + polymerase complex (TEC)
Supramolecule | Name: CPV VP4 + polymerase complex (TEC) / type: sample / ID: 1000 / Number unique components: 2 |
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-Macromolecule #1: VP4
Macromolecule | Name: VP4 / type: protein_or_peptide / ID: 1 / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: ![]() ![]() |
-Macromolecule #2: RNA polymerase
Macromolecule | Name: RNA polymerase / type: protein_or_peptide / ID: 2 / Recombinant expression: No / Database: NCBI |
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Source (natural) | Organism: ![]() ![]() |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 8 / Details: 70 mM Tris-Cl, 10 mM MgCl2, 100 mM NaCl, 2 mM GTP |
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Grid | Details: 200 mesh Quantifoil holey carbon film |
Vitrification | Cryogen name: ETHANE / Instrument: FEI VITROBOT MARK II |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Temperature | Average: 80 K |
Alignment procedure | Legacy - Electron beam tilt params: 0 |
Specialist optics | Energy filter - Name: Gatan Quantum Energy Filter |
Date | Mar 7, 2015 |
Image recording | Category: CCD / Film or detector model: GATAN K2 (4k x 4k) / Digitization - Sampling interval: 5 µm / Number real images: 4907 / Average electron dose: 40 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 36765 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.4 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 105000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
CTF correction | Details: Each particle |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 4.0 Å / Resolution method: OTHER / Software - Name: Frealign / Number images used: 81887 |
Final two d classification | Number classes: 1 |