|Entry||Database: PDB / ID: 6nur|
|Title||SARS-Coronavirus NSP12 bound to NSP7 and NSP8 co-factors|
|Keywords||VIRAL PROTEIN / coronavirus / polymerase / non-structural protein|
|Function / homology|
Function and homology information
modulation by virus of host autophagy / positive regulation of ubiquitin-specific protease activity / suppression by virus of host translation / mRNA methylation / RNA phosphodiester bond hydrolysis, exonucleolytic / Lys48-specific deubiquitinase activity / SARS coronavirus main proteinase / induction by virus of catabolism of host mRNA / host cell endoplasmic reticulum-Golgi intermediate compartment / suppression by virus of host ISG15 activity ...modulation by virus of host autophagy / positive regulation of ubiquitin-specific protease activity / suppression by virus of host translation / mRNA methylation / RNA phosphodiester bond hydrolysis, exonucleolytic / Lys48-specific deubiquitinase activity / SARS coronavirus main proteinase / induction by virus of catabolism of host mRNA / host cell endoplasmic reticulum-Golgi intermediate compartment / suppression by virus of host ISG15 activity / suppression by virus of host NF-kappaB transcription factor activity / cytoplasmic viral factory / 3'-5'-exoribonuclease activity / Hydrolases, Acting on ester bonds, Exoribonucleases producing 5'-phosphomonoesters / modulation by virus of host protein ubiquitination / omega peptidase activity / suppression by virus of host IRF3 activity / transcription, RNA-templated / 7-methylguanosine mRNA capping / viral transcription / positive stranded viral RNA replication / viral genome replication / Transferases, Transferring one-carbon groups, Methyltransferases / mRNA (nucleoside-2'-O-)-methyltransferase activity / mRNA (guanine-N7-)-methyltransferase activity / thiol-dependent ubiquitinyl hydrolase activity / helicase activity / DNA helicase / ubiquitinyl hydrolase 1 / single-stranded RNA binding / DNA helicase activity / methyltransferase activity / Hydrolases, Acting on peptide bonds (peptidases), Cysteine endopeptidases / host cell perinuclear region of cytoplasm / host cell membrane / viral protein processing / suppression by virus of host type I interferon-mediated signaling pathway / double-stranded RNA binding / RNA helicase / RNA-directed RNA polymerase / induction by virus of host autophagy / viral RNA genome replication / RNA helicase activity / cysteine-type endopeptidase activity / endonuclease activity / RNA-directed 5'-3' RNA polymerase activity / Hydrolases, Acting on ester bonds / transcription, DNA-templated / RNA binding / zinc ion binding / integral component of membrane / ATP binding / identical protein binding
Peptidase S1, PA clan / Papain-like viral protease / Endoribonuclease EndoU-like / Coronavirus NSP4, C-terminal domain superfamily / Replicase polyprotein 1a/1ab / NSP1 domain superfamily / Macro domain / RNA-directed RNA polymerase, catalytic domain / Peptidase C30, Coronavirus endopeptidase / NSP8 replicase superfamily ...Peptidase S1, PA clan / Papain-like viral protease / Endoribonuclease EndoU-like / Coronavirus NSP4, C-terminal domain superfamily / Replicase polyprotein 1a/1ab / NSP1 domain superfamily / Macro domain / RNA-directed RNA polymerase, catalytic domain / Peptidase C30, Coronavirus endopeptidase / NSP8 replicase superfamily / Coronavirus non-structural protein NSP16 / NSP11 / RNA polymerase, N-terminal, coronaviral / Peptidase C30/C16 / NSP9 replicase / Non structural protein 7 / Nsp3, coronavirus single-stranded poly(A) binding domain superfamily / NSP8 replicase / RNA synthesis protein NSP10, coronavirus / Non structural protein NSP1 / SARS coronavirus, polyprotein cleavage domain PL2pro / SARS coronavirus, non-structural protein 3, N-terminal / Non-structural protein 3, coronavirus single-stranded poly(A) binding domain / (+) RNA virus helicase core domain / Coronaviridae zinc-binding domain / P-loop containing nucleoside triphosphate hydrolase / S-adenosyl-L-methionine-dependent methyltransferase / Coronavirus nonstructural protein 4, C-terminal / Replicase polyprotein, nucleic acid-binding domain (NAR) / Coronavirus RNA synthesis protein NSP10 superfamily / Replicase NSP9 superfamily / Coronavirus, polyprotein cleavage domain PL2pro superfamily / NSP7 superfamily / Nsp15, N-terminal domain / Coronavirus polyprotein cleavage domain / nsp9 replicase / Papain like viral protease / nsp7 replicase / nsp8 replicase / RNA synthesis protein NSP10 / Non structural protein Nsp1 / Single-stranded poly(A) binding domain / Protein of unknown function (DUF3655) / NSP11 / Nucleic acid-binding domain (NAR) / Coronavirus nonstructural protein 4 C-terminus / RdRp of positive ssRNA viruses catalytic domain profile. / Peptidase family C16 domain profile. / Macro domain profile. / Coronavirus main protease (M-pro) domain profile. / Coronaviridae zinc-binding (CV ZBD) domain profile. / (+)RNA virus helicase core domain profile. / Replicase polyprotein, nucleic acid-binding domain superfamily / Coronavirus RPol N-terminus / Coronavirus NSP16 / Coronavirus endopeptidase C30 / Macro domain
Replicase polyprotein 1a / Replicase polyprotein 1ab
|Biological species||Human SARS coronavirus|
|Method||ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å|
|Authors||Kirchdoerfer, R.N. / Ward, A.B.|
|Funding support|| United States, 2items |
|Citation||Journal: Nat Commun / Year: 2019|
Title: Structure of the SARS-CoV nsp12 polymerase bound to nsp7 and nsp8 co-factors.
Authors: Robert N Kirchdoerfer / Andrew B Ward /
Abstract: Recent history is punctuated by the emergence of highly pathogenic coronaviruses such as SARS- and MERS-CoV into human circulation. Upon infecting host cells, coronaviruses assemble a multi-subunit ...Recent history is punctuated by the emergence of highly pathogenic coronaviruses such as SARS- and MERS-CoV into human circulation. Upon infecting host cells, coronaviruses assemble a multi-subunit RNA-synthesis complex of viral non-structural proteins (nsp) responsible for the replication and transcription of the viral genome. Here, we present the 3.1 Å resolution structure of the SARS-CoV nsp12 polymerase bound to its essential co-factors, nsp7 and nsp8, using single particle cryo-electron microscopy. nsp12 possesses an architecture common to all viral polymerases as well as a large N-terminal extension containing a kinase-like fold and is bound by two nsp8 co-factors. This structure illuminates the assembly of the coronavirus core RNA-synthesis machinery, provides key insights into nsp12 polymerase catalysis and fidelity and acts as a template for the design of novel antiviral therapeutics.
SummaryFull reportAbout validation report
|Structure viewer||Molecule: |
Downloads & links
|#1: Protein/peptide|| |
Mass: 109277.031 Da / Num. of mol.: 1 / Fragment: UNP residues 4370-5300
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human SARS coronavirus / Gene: rep, 1a-1b / Plasmid: pFastBac / Cell line (production host): Sf21 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P0C6X7
Mass: 21887.990 Da / Num. of mol.: 2 / Fragment: UNP residues 3920-4117
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human SARS coronavirus / Gene: 1a / Plasmid: pET46 / Production host: Escherichia coli BL21 (bacteria) / Strain (production host): BL21 Rosetta2 pLysS / References: UniProt: P0C6U8
Mass: 9333.869 Da / Num. of mol.: 1 / Fragment: UNP residues 3837-3919
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human SARS coronavirus / Gene: rep, 1a-1b / Plasmid: pET46 / Production host: Escherichia coli BL21 (bacteria) / Strain (production host): BL21 Rosetta2 pLysS / References: UniProt: P0C6X7
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction|
|Component||Name: SARS-Coronavirus NSP12 bound to NSP7 and NSP8 co-factors|
Type: COMPLEX / Entity ID: 1, 2, 3 / Source: RECOMBINANT
|Molecular weight||Value: 0.16 MDa / Experimental value: NO|
|Source (natural)||Organism: SARS coronavirus|
|Source (recombinant)||Organism: Spodoptera frugiperda (fall armyworm) / Cell: Sf21 / Plasmid: pFastBac|
|Buffer solution||pH: 7.4 |
Details: n-dodecyl-beta-D-maltopyranoside was added just prior to spotting samples onto holey EM grids.
|Specimen||Conc.: 3.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil, UltrAuFoil, R1.2/1.3|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K|
-Electron microscopy imaging
Model: Talos Arctica / Image courtesy: FEI Company
|Microscopy||Model: FEI TALOS ARCTICA|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy / Calibrated magnification: 43478 X / Calibrated defocus min: 400 nm / Calibrated defocus max: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE|
|Specimen holder||Cryogen: NITROGEN / Model: FEI TITAN KRIOS AUTOGRID HOLDER|
|Image recording||Average exposure time: 11.75 sec. / Electron dose: 50.5 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 1677|
|Image scans||Width: 3838 / Height: 3710 / Movie frames/image: 47 / Used frames/image: 1-47|
|CTF correction||Type: PHASE FLIPPING AND AMPLITUDE CORRECTION|
|Particle selection||Num. of particles selected: 2003890|
|Symmetry||Point symmetry: C1 (asymmetric)|
|3D reconstruction||Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 71046 / Symmetry type: POINT|
|Atomic model building||Space: REAL|
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