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- EMDB-15578: CryoEM structure of the Chikungunya virus nsP1 capping pores in c... -
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Open data
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Basic information
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Title | CryoEM structure of the Chikungunya virus nsP1 capping pores in covalent complex with a 7GMP cap structure | |||||||||
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Function / homology | ![]() host cell filopodium / ADP-ribose 1''-phosphate phosphatase / mRNA methyltransferase activity / mRNA 5'-triphosphate monophosphatase activity / mRNA 5'-phosphatase / polynucleotide 5'-phosphatase activity / polynucleotide adenylyltransferase / poly(A) RNA polymerase activity / 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 / polynucleotide adenylyltransferase / poly(A) RNA polymerase activity / 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 / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / RNA helicase activity / 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 | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.2 Å | |||||||||
![]() | Jones R / Hons M / Reguera J | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural basis and dynamics of Chikungunya alphavirus RNA capping by nsP1 capping pores. Authors: Rhian Jones / Michael Hons / Nadia Rabah / Noelia Zamarreño / Rocío Arranz / Juan Reguera / ![]() ![]() Abstract: Alphaviruses are emerging positive-stranded RNA viruses which replicate and transcribe their genomes in membranous organelles formed in the cell cytoplasm. The nonstructural protein 1 (nsP1) is ...Alphaviruses are emerging positive-stranded RNA viruses which replicate and transcribe their genomes in membranous organelles formed in the cell cytoplasm. The nonstructural protein 1 (nsP1) is responsible for viral RNA capping and gates the replication organelles by assembling into monotopic membrane-associated dodecameric pores. The capping pathway is unique to Alphaviruses; beginning with the N methylation of a guanosine triphosphate (GTP) molecule, followed by the covalent linkage of an mGMP group to a conserved histidine in nsP1 and the transfer of this cap structure to a diphosphate RNA. Here, we provide structural snapshots of different stages of the reaction pathway showing how nsP1 pores recognize the substrates of the methyl-transfer reaction, GTP and S-adenosyl methionine (SAM), how the enzyme reaches a metastable postmethylation state with SAH and mGTP in the active site, and the subsequent covalent transfer of mGMP to nsP1 triggered by the presence of RNA and postdecapping reaction conformational changes inducing the opening of the pore. In addition, we biochemically characterize the capping reaction, demonstrating specificity for the RNA substrate and the reversibility of the cap transfer resulting in decapping activity and the release of reaction intermediates. Our data identify the molecular determinants allowing each pathway transition, providing an explanation for the need for the SAM methyl donor all along the pathway and clues about the conformational rearrangements associated to the enzymatic activity of nsP1. Together, our results set ground for the structural and functional understanding of alphavirus RNA-capping and the design of antivirals. | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 157.7 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 18.9 KB 18.9 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 12.8 KB | Display | ![]() |
Images | ![]() | 97.1 KB | ||
Others | ![]() ![]() | 164.9 MB 165 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 827.2 KB | Display | ![]() |
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Full document | ![]() | 826.7 KB | Display | |
Data in XML | ![]() | 20.5 KB | Display | |
Data in CIF | ![]() | 26.8 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8apxMC ![]() 8aovC ![]() 8aowC ![]() 8aoxC ![]() 8axvC M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.855 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_15578_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_15578_half_map_2.map | ||||||||||||
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Density Histograms |
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Sample components
-Entire : CHIKV nsP1 capping pores covalently linked to m7GMP cap via H37
Entire | Name: CHIKV nsP1 capping pores covalently linked to m7GMP cap via H37 |
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Components |
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-Supramolecule #1: CHIKV nsP1 capping pores covalently linked to m7GMP cap via H37
Supramolecule | Name: CHIKV nsP1 capping pores covalently linked to m7GMP cap via H37 type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 720 KDa |
-Macromolecule #1: Polyprotein P1234
Macromolecule | Name: Polyprotein P1234 / type: protein_or_peptide / ID: 1 / Number of copies: 12 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 53.011754 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MDPVYVDIDA DSAFLKALQR AYPMFEVEPR QVTPND(PJ3)ANA RAFSHLAIKL IEQEIDPDST ILDIGSAPAR RMMSDR KYH CVCPMRSAED PERLANYARK LASAAGKVLD RNISGKIGDL QAVMAVPDTE TPTFCLHTDV SCRQRADVAI YQDVYAV HA PTSLYHQAIK ...String: MDPVYVDIDA DSAFLKALQR AYPMFEVEPR QVTPND(PJ3)ANA RAFSHLAIKL IEQEIDPDST ILDIGSAPAR RMMSDR KYH CVCPMRSAED PERLANYARK LASAAGKVLD RNISGKIGDL QAVMAVPDTE TPTFCLHTDV SCRQRADVAI YQDVYAV HA PTSLYHQAIK GVRLAYWVGF DTTPFMYNAM AGAYPSYSTN WADEQVLKAK NIGLCSTDLT EGRRGKLSIM RGKKLEPC D RVLFSVGSTL YPESRKLLKS WHLPSVFHLK GKLSFTCRCD TVVSCEGYVV KRITMSPGLY GKTTGYAVTH HADGFLMCK TTDTVDGERV SFSVCTYVPA TICDQMTGIL ATEVTPEDAQ KLLVGLNQRI VVNGRTQRNT NTMKNYMIPV VAQAFSKWAK ECRKDMEDE KLLGVRERTL TCCCLWAFKK QKTHTVYKRP DTQSIQKVQA EFDSFVVPSL WSSGLSIPLR TRIK |
-Macromolecule #2: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 2 / Number of copies: 12 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Macromolecule #3: S-ADENOSYL-L-HOMOCYSTEINE
Macromolecule | Name: S-ADENOSYL-L-HOMOCYSTEINE / type: ligand / ID: 3 / Number of copies: 12 / Formula: SAH |
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Molecular weight | Theoretical: 384.411 Da |
Chemical component information | ![]() ChemComp-SAH: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.2 mg/mL | ||||||||||||||||||
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Buffer | pH: 7.6 Component:
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Grid | Model: Quantifoil R2/2 / Material: GOLD / Mesh: 300 / Support film - Material: GRAPHENE OXIDE / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 10 sec. / Pretreatment - Atmosphere: AIR | ||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV | ||||||||||||||||||
Details | Sample incubated with 10x molar excess of m7GTP, SAH and 27mer CHIKV RNA to form the complex prior to freezing in the presence of magnesium. |
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Electron microscopy
Microscope | TFS TALOS |
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Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 773 / Average exposure time: 38.0 sec. / Average electron dose: 32.0 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm |
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Image processing
-Atomic model buiding 1
Initial model | Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Target criteria: Correlation coefficient |
Output model | ![]() PDB-8apx: |