+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-11007 | |||||||||||||||||||||
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Title | Structure of replicating SARS-CoV-2 polymerase | |||||||||||||||||||||
Map data | Main composite map used in refinement. | |||||||||||||||||||||
Sample |
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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 / snRNP Assembly / TRAF3-dependent IRF activation pathway / Replication of the SARS-CoV-2 genome / double membrane vesicle viral factory outer membrane / Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters / SARS coronavirus main proteinase / host cell endosome / 3'-5'-RNA exonuclease activity / host cell endoplasmic reticulum-Golgi intermediate compartment / : / 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 / SARS-CoV-2 modulates host translation machinery / omega peptidase activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of IRF3 activity / mRNA (guanine-N7)-methyltransferase / host cell Golgi apparatus / methyltransferase cap1 / symbiont-mediated perturbation of host ubiquitin-like protein modification / mRNA (nucleoside-2'-O-)-methyltransferase activity / DNA helicase / 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 endoplasmic reticulum membrane / host cell perinuclear region of cytoplasm / viral protein processing / lyase activity / RNA helicase / induction by virus of host autophagy / RNA-directed RNA polymerase / copper ion binding / symbiont-mediated suppression of host gene expression / viral RNA genome replication / cysteine-type endopeptidase activity / RNA-dependent RNA polymerase activity / DNA-templated transcription / lipid binding / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / 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 / synthetic construct (others) | |||||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.9 Å | |||||||||||||||||||||
Authors | Hillen HS / Kokic G / Farnung L / Dienemann C / Tegunov D / Cramer P | |||||||||||||||||||||
Funding support | Germany, 6 items
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Citation | Journal: Nature / Year: 2020 Title: Structure of replicating SARS-CoV-2 polymerase. Authors: Hauke S Hillen / Goran Kokic / Lucas Farnung / Christian Dienemann / Dimitry Tegunov / Patrick Cramer / Abstract: The new coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) uses an RNA-dependent RNA polymerase (RdRp) for the replication of its genome and the transcription of its genes. ...The new coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) uses an RNA-dependent RNA polymerase (RdRp) for the replication of its genome and the transcription of its genes. Here we present a cryo-electron microscopy structure of the SARS-CoV-2 RdRp in an active form that mimics the replicating enzyme. The structure comprises the viral proteins non-structural protein 12 (nsp12), nsp8 and nsp7, and more than two turns of RNA template-product duplex. The active-site cleft of nsp12 binds to the first turn of RNA and mediates RdRp activity with conserved residues. Two copies of nsp8 bind to opposite sides of the cleft and position the second turn of RNA. Long helical extensions in nsp8 protrude along exiting RNA, forming positively charged 'sliding poles'. These sliding poles can account for the known processivity of RdRp that is required for replicating the long genome of coronaviruses. Our results enable a detailed analysis of the inhibitory mechanisms that underlie the antiviral activity of substances such as remdesivir, a drug for the treatment of coronavirus disease 2019 (COVID-19). | |||||||||||||||||||||
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_11007.map.gz | 49.2 MB | EMDB map data format | |
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Header (meta data) | emd-11007-v30.xml emd-11007.xml | 32.7 KB 32.7 KB | Display Display | EMDB header |
Images | emd_11007.png | 135.7 KB | ||
Masks | emd_11007_msk_1.map emd_11007_msk_2.map | 64 MB 52.7 MB | Mask map | |
Others | emd_11007_additional_1.map.gz emd_11007_additional_2.map.gz emd_11007_additional_3.map.gz emd_11007_half_map_1.map.gz emd_11007_half_map_2.map.gz | 49.5 MB 49.5 MB 49.6 MB 40.8 MB 40.8 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-11007 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-11007 | HTTPS FTP |
-Related structure data
Related structure data | 6yytMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | |
EM raw data | EMPIAR-10409 (Title: Structure of replicating SARS-CoV-2 polymerase / Data size: 2.5 TB Data #1: Unaligned TIF movies of SARS-CoV2 RdRp in complex with nsp7, nsp8 and RNA (part 1) [micrographs - multiframe] Data #2: Unaligned TIF movies of SARS-CoV2 RdRp in complex with nsp7, nsp8 and RNA (part 2) [micrographs - multiframe] Data #3: Warp picked and extracted particles [picked particles - single frame - processed] Data #4: Reextraced particles at 1.3 A/px (for Map2 and Map3) [picked particles - single frame - processed] Data #5: Refined particles (Map 1) [picked particles - single frame - processed] Data #6: Refined particles (Map 2) [picked particles - single frame - processed] Data #7: Refined particles (Map 3) [picked particles - single frame - processed]) |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_11007.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Main composite map used in refinement. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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_11007_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Mask #2
File | emd_11007_msk_2.map | ||||||||||||
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Density Histograms |
-Additional map: Map2 nsp8b focus half map 2.
File | emd_11007_additional_1.map | ||||||||||||
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Annotation | Map2_nsp8b focus half map 2. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: Map2 nsp8b focus half map 1.
File | emd_11007_additional_2.map | ||||||||||||
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Annotation | Map2_nsp8b focus half map 1. | ||||||||||||
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Density Histograms |
-Additional map: Map2 nsp8b class half map 1.
File | emd_11007_additional_3.map | ||||||||||||
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Annotation | Map2_nsp8b _class half map 1. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Map1 core half map 2.
File | emd_11007_half_map_1.map | ||||||||||||
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Annotation | Map1_core half map 2. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Map1 core half map 1.
File | emd_11007_half_map_2.map | ||||||||||||
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Annotation | Map1_core half map 1. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : RNA-dependent RNA polymerase complex consisting of nsp12, nsp8, n...
Entire | Name: RNA-dependent RNA polymerase complex consisting of nsp12, nsp8, nsp7 and duplex RNA. |
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Components |
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-Supramolecule #1: RNA-dependent RNA polymerase complex consisting of nsp12, nsp8, n...
Supramolecule | Name: RNA-dependent RNA polymerase complex consisting of nsp12, nsp8, nsp7 and duplex RNA. type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4 |
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Molecular weight | Theoretical: 186 KDa |
-Supramolecule #2: nsp12
Supramolecule | Name: nsp12 / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Recombinant expression | Organism: Spodoptera aff. frugiperda 1 BOLD-2017 (butterflies/moths) |
-Supramolecule #3: duplex RNA
Supramolecule | Name: duplex RNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4 |
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Source (natural) | Organism: synthetic construct (others) |
Recombinant expression | Organism: synthetic construct (others) |
-Supramolecule #4: nsp8, nsp7
Supramolecule | Name: nsp8, nsp7 / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #2-#3 |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
-Macromolecule #1: nsp12
Macromolecule | Name: nsp12 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: ubiquitinyl hydrolase 1 |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Molecular weight | Theoretical: 107.053227 KDa |
Recombinant expression | Organism: Spodoptera aff. frugiperda 1 BOLD-2017 (butterflies/moths) |
Sequence | String: SNASADAQSF LNRVCGVSAA RLTPCGTGTS TDVVYRAFDI YNDKVAGFAK FLKTNCCRFQ EKDEDDNLID SYFVVKRHTF SNYQHEETI YNLLKDCPAV AKHDFFKFRI DGDMVPHISR QRLTKYTMAD LVYALRHFDE GNCDTLKEIL VTYNCCDDDY F NKKDWYDF ...String: SNASADAQSF LNRVCGVSAA RLTPCGTGTS TDVVYRAFDI YNDKVAGFAK FLKTNCCRFQ EKDEDDNLID SYFVVKRHTF SNYQHEETI YNLLKDCPAV AKHDFFKFRI DGDMVPHISR QRLTKYTMAD LVYALRHFDE GNCDTLKEIL VTYNCCDDDY F NKKDWYDF VENPDILRVY ANLGERVRQA LLKTVQFCDA MRNAGIVGVL TLDNQDLNGN WYDFGDFIQT TPGSGVPVVD SY YSLLMPI LTLTRALTAE SHVDTDLTKP YIKWDLLKYD FTEERLKLFD RYFKYWDQTY HPNCVNCLDD RCILHCANFN VLF STVFPP TSFGPLVRKI FVDGVPFVVS TGYHFRELGV VHNQDVNLHS SRLSFKELLV YAADPAMHAA SGNLLLDKRT TCFS VAALT NNVAFQTVKP GNFNKDFYDF AVSKGFFKEG SSVELKHFFF AQDGNAAISD YDYYRYNLPT MCDIRQLLFV VEVVD KYFD CYDGGCINAN QVIVNNLDKS AGFPFNKWGK ARLYYDSMSY EDQDALFAYT KRNVIPTITQ MNLKYAISAK NRARTV AGV SICSTMTNRQ FHQKLLKSIA ATRGATVVIG TSKFYGGWHN MLKTVYSDVE NPHLMGWDYP KCDRAMPNML RIMASLV LA RKHTTCCSLS HRFYRLANEC AQVLSEMVMC GGSLYVKPGG TSSGDATTAY ANSVFNICQA VTANVNALLS TDGNKIAD K YVRNLQHRLY ECLYRNRDVD TDFVNEFYAY LRKHFSMMIL SDDAVVCFNS TYASQGLVAS IKNFKSVLYY QNNVFMSEA KCWTETDLTK GPHEFCSQHT MLVKQGDDYV YLPYPDPSRI LGAGCFVDDI VKTDGTLMIE RFVSLAIDAY PLTKHPNQEY ADVFHLYLQ YIRKLHDELT GHMLDMYSVM LTNDNTSRYW EPEFYEAMYT PHTVLQ |
-Macromolecule #2: nsp8
Macromolecule | Name: nsp8 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO / EC number: ubiquitinyl hydrolase 1 |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Molecular weight | Theoretical: 22.175309 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: SNAAIASEFS SLPSYAAFAT AQEAYEQAVA NGDSEVVLKK LKKSLNVAKS EFDRDAAMQR KLEKMADQAM TQMYKQARSE DKRAKVTSA MQTMLFTMLR KLDNDALNNI INNARDGCVP LNIIPLTTAA KLMVVIPDYN TYKNTCDGTT FTYASALWEI Q QVVDADSK ...String: SNAAIASEFS SLPSYAAFAT AQEAYEQAVA NGDSEVVLKK LKKSLNVAKS EFDRDAAMQR KLEKMADQAM TQMYKQARSE DKRAKVTSA MQTMLFTMLR KLDNDALNNI INNARDGCVP LNIIPLTTAA KLMVVIPDYN TYKNTCDGTT FTYASALWEI Q QVVDADSK IVQLSEISMD NSPNLAWPLI VTALRANSAV KLQ |
-Macromolecule #3: nsp7
Macromolecule | Name: nsp7 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: ubiquitinyl hydrolase 1 |
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Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 |
Molecular weight | Theoretical: 9.521062 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: SNASKMSDVK CTSVVLLSVL QQLRVESSSK LWAQCVQLHN DILLAKDTTE AFEKMVSLLS VLLSMQGAVD INKLCEEMLD NRATLQ |
-Macromolecule #4: RNA product
Macromolecule | Name: RNA product / type: rna / ID: 4 / Number of copies: 4 |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 5.687372 KDa |
Sequence | String: UUUUCAUGCU ACGCGUAG |
-Macromolecule #5: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 5 / Number of copies: 2 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.4 Component:
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Grid | Model: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.4 kPa / Details: 15 mA, Pelco EasyGlow | |||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Calibrated magnification: 105000 / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 105000 |
Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 8168 / Average exposure time: 2.2 sec. / Average electron dose: 60.0 e/Å2 Details: Images were collected at 30 degree stage tilt. Non-superres. |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Particle selection | Number selected: 1300000 |
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CTF correction | Software - Name: Warp (ver. 1.07 BETA) |
Startup model | Type of model: PDB ENTRY PDB model - PDB ID: |
Initial angle assignment | Type: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 2.14.2) |
Final 3D classification | Number classes: 2 / Software - Name: RELION (ver. 3.0.7) |
Final angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0.7) |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0.7) / Number images used: 418000 |