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Yorodumi- EMDB-11994: Structure of elongating SARS-CoV-2 RNA-dependent RNA polymerase w... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-11994 | ||||||||||||||||||||||||
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Title | Structure of elongating SARS-CoV-2 RNA-dependent RNA polymerase with Remdesivir at position -4 (structure 2) | ||||||||||||||||||||||||
Map data | Denoised main map. | ||||||||||||||||||||||||
Sample |
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Keywords | SARS-CoV-2 / Polymerase / Transcription / Replication / RNA / Remdesivir / Inhibition / Nucleotide Analogue / VIRAL PROTEIN | ||||||||||||||||||||||||
Function / homology | Function and homology information protein guanylyltransferase activity / RNA endonuclease activity, producing 3'-phosphomonoesters / mRNA guanylyltransferase activity / 5'-3' RNA helicase activity / Lyases; Phosphorus-oxygen lyases / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of TBK1 activity / Assembly of the SARS-CoV-2 Replication-Transcription Complex (RTC) / Maturation of replicase proteins / ISG15-specific peptidase activity / Transcription of SARS-CoV-2 sgRNAs ...protein guanylyltransferase activity / RNA endonuclease activity, producing 3'-phosphomonoesters / mRNA guanylyltransferase activity / 5'-3' RNA helicase activity / Lyases; Phosphorus-oxygen lyases / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of TBK1 activity / Assembly of the SARS-CoV-2 Replication-Transcription Complex (RTC) / Maturation of replicase proteins / ISG15-specific peptidase activity / Transcription of SARS-CoV-2 sgRNAs / Translation of Replicase and Assembly of the Replication Transcription Complex / TRAF3-dependent IRF activation pathway / Replication of the SARS-CoV-2 genome / snRNP Assembly / double membrane vesicle viral factory outer membrane / Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters / 5'-3' DNA helicase activity / SARS coronavirus main proteinase / 3'-5'-RNA exonuclease activity / host cell endoplasmic reticulum-Golgi intermediate compartment / host cell endosome / symbiont-mediated suppression of host toll-like receptor signaling pathway / symbiont-mediated degradation of host mRNA / mRNA guanylyltransferase / symbiont-mediated suppression of host ISG15-protein conjugation / G-quadruplex RNA binding / omega peptidase activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of IRF3 activity / SARS-CoV-2 modulates host translation machinery / mRNA (guanine-N7)-methyltransferase / host cell Golgi apparatus / methyltransferase cap1 / symbiont-mediated perturbation of host ubiquitin-like protein modification / DNA helicase / mRNA (nucleoside-2'-O-)-methyltransferase activity / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / ubiquitinyl hydrolase 1 / cysteine-type deubiquitinase activity / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / single-stranded RNA binding / host cell perinuclear region of cytoplasm / host cell endoplasmic reticulum membrane / viral protein processing / lyase activity / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / RNA helicase / induction by virus of host autophagy / copper ion binding / cysteine-type endopeptidase activity / RNA-directed RNA polymerase / viral RNA genome replication / virus-mediated perturbation of host defense response / RNA-dependent RNA polymerase activity / DNA-templated transcription / lipid binding / host cell nucleus / SARS-CoV-2 activates/modulates innate and adaptive immune responses / ATP hydrolysis activity / proteolysis / RNA binding / zinc ion binding / ATP binding / membrane Similarity search - Function | ||||||||||||||||||||||||
Biological species | Severe acute respiratory syndrome coronavirus 2 | ||||||||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||||||||||||||||||||
Authors | Kokic G / Hillen HS | ||||||||||||||||||||||||
Funding support | Germany, 7 items
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Citation | Journal: Nat Commun / Year: 2021 Title: Mechanism of SARS-CoV-2 polymerase stalling by remdesivir. Authors: Goran Kokic / Hauke S Hillen / Dimitry Tegunov / Christian Dienemann / Florian Seitz / Jana Schmitzova / Lucas Farnung / Aaron Siewert / Claudia Höbartner / Patrick Cramer / Abstract: Remdesivir is the only FDA-approved drug for the treatment of COVID-19 patients. The active form of remdesivir acts as a nucleoside analog and inhibits the RNA-dependent RNA polymerase (RdRp) of ...Remdesivir is the only FDA-approved drug for the treatment of COVID-19 patients. The active form of remdesivir acts as a nucleoside analog and inhibits the RNA-dependent RNA polymerase (RdRp) of coronaviruses including SARS-CoV-2. Remdesivir is incorporated by the RdRp into the growing RNA product and allows for addition of three more nucleotides before RNA synthesis stalls. Here we use synthetic RNA chemistry, biochemistry and cryo-electron microscopy to establish the molecular mechanism of remdesivir-induced RdRp stalling. We show that addition of the fourth nucleotide following remdesivir incorporation into the RNA product is impaired by a barrier to further RNA translocation. This translocation barrier causes retention of the RNA 3'-nucleotide in the substrate-binding site of the RdRp and interferes with entry of the next nucleoside triphosphate, thereby stalling RdRp. In the structure of the remdesivir-stalled state, the 3'-nucleotide of the RNA product is matched and located with the template base in the active center, and this may impair proofreading by the viral 3'-exonuclease. These mechanistic insights should facilitate the quest for improved antivirals that target coronavirus replication. | ||||||||||||||||||||||||
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_11994.map.gz | 48 MB | EMDB map data format | |
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Header (meta data) | emd-11994-v30.xml emd-11994.xml | 25.8 KB 25.8 KB | Display Display | EMDB header |
Images | emd_11994.png | 132.8 KB | ||
Masks | emd_11994_msk_1.map | 13.9 MB | Mask map | |
Filedesc metadata | emd-11994.cif.gz | 7.6 KB | ||
Others | emd_11994_half_map_1.map.gz emd_11994_half_map_2.map.gz | 7.1 MB 7.1 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-11994 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-11994 | HTTPS FTP |
-Validation report
Summary document | emd_11994_validation.pdf.gz | 765.5 KB | Display | EMDB validaton report |
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Full document | emd_11994_full_validation.pdf.gz | 765.1 KB | Display | |
Data in XML | emd_11994_validation.xml.gz | 9.9 KB | Display | |
Data in CIF | emd_11994_validation.cif.gz | 11.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-11994 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-11994 | HTTPS FTP |
-Related structure data
Related structure data | 7b3cMC 7b3bC 7b3dC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_11994.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Denoised main map. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.834 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_11994_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: Half map 1
File | emd_11994_half_map_1.map | ||||||||||||
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Annotation | Half map 1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 2
File | emd_11994_half_map_2.map | ||||||||||||
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Annotation | Half map 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : SARS-CoV-2 RNA-dependent RNA polymerase complex (nsp12, nsp7, nsp...
+Supramolecule #1: SARS-CoV-2 RNA-dependent RNA polymerase complex (nsp12, nsp7, nsp...
+Supramolecule #2: SARS-CoV-2 RNA-dependent RNA polymerase (nsp12)
+Supramolecule #3: SARS-CoV-2 nsp7 and 8
+Supramolecule #4: DNA and RNA
+Macromolecule #1: RNA-directed RNA polymerase nsp12
+Macromolecule #2: Non-structural protein 8
+Macromolecule #3: Non-structural protein 7
+Macromolecule #4: DNA/RNA (5'-R(P*CP*UP*AP*CP*GP*CP*A)-D(P*(RMP))-R(P*GP*UP*G)-3')
+Macromolecule #5: RNA (5'-R(P*UP*CP*AP*CP*UP*UP*GP*CP*GP*UP*AP*G)-3')
+Macromolecule #6: ZINC ION
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 2 mg/mL | ||||||||||
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Buffer | pH: 7.4 Component:
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Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Details | 30 deg tilt. |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 60.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Calibrated defocus max: 0.0017 µm / Calibrated defocus min: 0.0004 µm / Illumination mode: OTHER / Imaging mode: OTHER / Cs: 2.7 mm / Nominal magnification: 105000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |