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Yorodumi- EMDB-33591: Molecular architecture of the chikungunya virus replication complex -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-33591 | |||||||||
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Title | Molecular architecture of the chikungunya virus replication complex | |||||||||
Map data | main map | |||||||||
Sample |
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Keywords | replicase / replication complex / viral RNA replication / capping / helicase / protease / RNA-dependent RNA polymerase / RdRp / VIRAL PROTEIN / VIRAL PROTEIN-RNA complex | |||||||||
Function / homology | Function and homology information host cell filopodium / ADP-ribose 1''-phosphate phosphatase / mRNA methyltransferase activity / mRNA 5'-triphosphate monophosphatase activity / mRNA 5'-phosphatase / polynucleotide 5'-phosphatase activity / poly(A) RNA polymerase activity / polynucleotide adenylyltransferase / regulation of cytoskeleton organization / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity ...host cell filopodium / ADP-ribose 1''-phosphate phosphatase / mRNA methyltransferase activity / mRNA 5'-triphosphate monophosphatase activity / mRNA 5'-phosphatase / polynucleotide 5'-phosphatase activity / poly(A) RNA polymerase activity / polynucleotide adenylyltransferase / regulation of cytoskeleton organization / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / 7-methylguanosine mRNA capping / cysteine-type peptidase activity / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT1 activity / Transferases; Transferring one-carbon groups; Methyltransferases / host cell cytoplasmic vesicle membrane / Transferases; Transferring phosphorus-containing groups; Nucleotidyltransferases / nucleoside-triphosphate phosphatase / methylation / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / RNA helicase activity / symbiont-mediated suppression of host innate immune response / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / RNA helicase / RNA-directed RNA polymerase / viral RNA genome replication / RNA-dependent RNA polymerase activity / virus-mediated perturbation of host defense response / DNA-templated transcription / host cell nucleus / GTP binding / host cell plasma membrane / ATP hydrolysis activity / proteolysis / RNA binding / ATP binding / membrane / metal ion binding Similarity search - Function | |||||||||
Biological species | Chikungunya virus strain S27-African prototype / Onyong-nyong virus / in vitro transcription vector pT7-TP(deltai) (others) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.8 Å | |||||||||
Authors | Tan YB / Luo D | |||||||||
Funding support | Singapore, 1 items
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Citation | Journal: Sci Adv / Year: 2022 Title: Molecular architecture of the Chikungunya virus replication complex. Authors: Yaw Bia Tan / David Chmielewski / Michelle Cheok Yien Law / Kuo Zhang / Yu He / Muyuan Chen / Jing Jin / Dahai Luo / Abstract: To better understand how positive-strand (+) RNA viruses assemble membrane-associated replication complexes (RCs) to synthesize, process, and transport viral RNA in virus-infected cells, we ...To better understand how positive-strand (+) RNA viruses assemble membrane-associated replication complexes (RCs) to synthesize, process, and transport viral RNA in virus-infected cells, we determined both the high-resolution structure of the core RNA replicase of chikungunya virus and the native RC architecture in its cellular context at subnanometer resolution, using in vitro reconstitution and in situ electron cryotomography, respectively. Within the core RNA replicase, the viral polymerase nsP4, which is in complex with nsP2 helicase-protease, sits in the central pore of the membrane-anchored nsP1 RNA-capping ring. The addition of a large cytoplasmic ring next to the C terminus of nsP1 forms the holo-RNA-RC as observed at the neck of spherules formed in virus-infected cells. These results represent a major conceptual advance in elucidating the molecular mechanisms of RNA virus replication and the principles underlying the molecular architecture of RCs, likely to be shared with many pathogenic (+) RNA viruses. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_33591.map.gz | 123.2 MB | EMDB map data format | |
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Header (meta data) | emd-33591-v30.xml emd-33591.xml | 32.1 KB 32.1 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_33591_fsc.xml | 13.1 KB | Display | FSC data file |
Images | emd_33591.png | 224.3 KB | ||
Masks | emd_33591_msk_1.map | 244.1 MB | Mask map | |
Filedesc metadata | emd-33591.cif.gz | 9.4 KB | ||
Others | emd_33591_additional_1.map.gz emd_33591_half_map_1.map.gz emd_33591_half_map_2.map.gz | 230.2 MB 226.8 MB 226.8 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-33591 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-33591 | HTTPS FTP |
-Validation report
Summary document | emd_33591_validation.pdf.gz | 1.2 MB | Display | EMDB validaton report |
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Full document | emd_33591_full_validation.pdf.gz | 1.2 MB | Display | |
Data in XML | emd_33591_validation.xml.gz | 21.6 KB | Display | |
Data in CIF | emd_33591_validation.cif.gz | 28 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-33591 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-33591 | HTTPS FTP |
-Related structure data
Related structure data | 7y38MC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_33591.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Annotation | main map | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.1 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_33591_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: sharpen map
File | emd_33591_additional_1.map | ||||||||||||
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Annotation | sharpen map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: halfmap B (coordinated to the same as main map)
File | emd_33591_half_map_1.map | ||||||||||||
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Annotation | halfmap B (coordinated to the same as main map) | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: halfmap A (coordinated to the same as main map)
File | emd_33591_half_map_2.map | ||||||||||||
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Annotation | halfmap A (coordinated to the same as main map) | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : Alphavirus Replication Complex
+Supramolecule #1: Alphavirus Replication Complex
+Supramolecule #2: nsP1 dodecameric ring
+Supramolecule #3: nsP4 RdRp; docked within nsP1 central cavity
+Supramolecule #4: nsP2 helicase-protease; docked on top of viral complex with synth...
+Macromolecule #1: mRNA-capping enzyme nsP1,affinity-tag (strepII-3XFLAG)
+Macromolecule #2: RNA-directed RNA polymerase nsP4
+Macromolecule #3: Protease nsP2
+Macromolecule #4: RNA (5'-R(P*CP*CP*A)-3')
+Macromolecule #5: ZINC ION
+Macromolecule #6: ADENOSINE-5'-TRIPHOSPHATE
+Macromolecule #7: GUANOSINE-5'-TRIPHOSPHATE
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 10 mg/mL | ||||||||||||||||||||||||
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Buffer | pH: 8 Component:
Details: 25mM HEPES, 75mM NaCl, 20mM KCl, 5mM MgOAc, 2.5mM MnCl2, 5mM TCEP, 1.25% Sucrose, pH8 | ||||||||||||||||||||||||
Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: GRAPHENE / Support film - topology: CONTINUOUS / Support film - Film thickness: 3.45 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 10 sec. / Pretreatment - Atmosphere: OTHER | ||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 273.15 K / Instrument: FEI VITROBOT MARK III / Details: wait 45s, blot 2s at force -2. | ||||||||||||||||||||||||
Details | The sample was in vitro reconstituted and purified via affinity pulldown and ion-exchanged. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Temperature | Min: 70.0 K / Max: 80.0 K |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Number grids imaged: 1 / Number real images: 7865 / Average exposure time: 5.0 sec. / Average electron dose: 34.0 e/Å2 / Details: movie-mode at 40 FPS |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.6 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Residue range: 1-516 / Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Details | ChimeraX was used to fit the 7DOP ring and built/refined using coot/phenix. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Overall B value: 63.66 / Target criteria: Correlation Coefficient |
Output model | PDB-7y38: |