+Open data
-Basic information
Entry | Database: PDB / ID: 8urb | ||||||||||||
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Title | Porcine epidemic diarrhea virus complete core polymerase complex | ||||||||||||
Components |
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Keywords | REPLICATION / TRANSFERASE/RNA / Coronavirus / nsp12 / RNA polymerase / PEDV / TRANSFERASE-RNA complex | ||||||||||||
Function / homology | Function and homology information host cell membrane / viral genome replication / host cell endoplasmic reticulum-Golgi intermediate compartment / 3'-5'-RNA exonuclease activity / 5'-3' DNA helicase activity / omega peptidase activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of IRF3 activity / symbiont-mediated perturbation of host ubiquitin-like protein modification / endonuclease activity / mRNA (nucleoside-2'-O-)-methyltransferase activity ...host cell membrane / viral genome replication / host cell endoplasmic reticulum-Golgi intermediate compartment / 3'-5'-RNA exonuclease activity / 5'-3' DNA helicase activity / omega peptidase activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of IRF3 activity / symbiont-mediated perturbation of host ubiquitin-like protein modification / endonuclease activity / mRNA (nucleoside-2'-O-)-methyltransferase activity / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / cysteine-type deubiquitinase activity / RNA helicase activity / host cell perinuclear region of cytoplasm / viral protein processing / lyase activity / induction by virus of host autophagy / viral translational frameshifting / cysteine-type endopeptidase activity / RNA-dependent RNA polymerase activity / virus-mediated perturbation of host defense response / DNA-templated transcription / proteolysis / RNA binding / zinc ion binding / ATP binding / membrane Similarity search - Function | ||||||||||||
Biological species | Porcine epidemic diarrhea virus synthetic construct (others) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||||||||
Authors | Anderson, T.K. / Kirchdoerfer, R.N. | ||||||||||||
Funding support | United States, 3items
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Citation | Journal: bioRxiv / Year: 2023 Title: An alphacoronavirus polymerase structure reveals conserved co-factor functions. Authors: Thomas K Anderson / Peter J Hoferle / Kenneth W Lee / Joshua J Coon / Robert N Kirchdoerfer / Abstract: Coronaviruses are a diverse subfamily of viruses containing pathogens of humans and animals. This subfamily of viruses replicates their RNA genomes using a core polymerase complex composed of viral ...Coronaviruses are a diverse subfamily of viruses containing pathogens of humans and animals. This subfamily of viruses replicates their RNA genomes using a core polymerase complex composed of viral non-structural proteins: nsp7, nsp8 and nsp12. Most of our understanding of coronavirus molecular biology comes from the betacoronaviruses like SARS-CoV and SARS-CoV-2, the latter of which is the causative agent of COVID-19. In contrast, members of the alphacoronavirus genus are relatively understudied despite their importance in human and animal health. Here we have used cryo-electron microscopy to determine the structure of the alphacoronavirus porcine epidemic diarrhea virus (PEDV) core polymerase complex bound to RNA. Our structure shows an unexpected nsp8 stoichiometry in comparison to other published coronavirus polymerase structures. Biochemical analysis shows that the N-terminal extension of one nsp8 is not required for RNA synthesis for alpha and betacoronaviruses as previously hypothesized. Our work shows the importance of studying diverse coronaviruses to reveal aspects of coronavirus replication while also identifying areas of conservation to be targeted by antiviral drugs. | ||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8urb.cif.gz | 543.6 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8urb.ent.gz | 445.4 KB | Display | PDB format |
PDBx/mmJSON format | 8urb.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8urb_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 8urb_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 8urb_validation.xml.gz | 48.5 KB | Display | |
Data in CIF | 8urb_validation.cif.gz | 73.8 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ur/8urb ftp://data.pdbj.org/pub/pdb/validation_reports/ur/8urb | HTTPS FTP |
-Related structure data
Related structure data | 42488MC 8g6rC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein , 3 types, 4 molecules ABDC
#1: Protein | Mass: 108994.758 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Porcine epidemic diarrhea virus / Gene: 1b, ORF1a / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: U6BRU0 | ||
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#2: Protein | Mass: 21716.838 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Porcine epidemic diarrhea virus / Gene: 1b, ORF1a / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: U6BRU0 #3: Protein | | Mass: 9230.511 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Porcine epidemic diarrhea virus / Gene: 1b, ORF1a / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: U6BRU0 |
-RNA chain , 2 types, 2 molecules IJ
#4: RNA chain | Mass: 10468.266 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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#5: RNA chain | Mass: 17740.572 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
-Non-polymers , 1 types, 2 molecules
#6: Chemical |
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-Details
Has ligand of interest | Y |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Core polymerase complex of the porcine epidemic diarrhea virus composed of nsp12, nsp7, nsp8, and a short RNA substrate Type: COMPLEX / Entity ID: #1-#5 / Source: RECOMBINANT | ||||||||||||||||||||||||||||||
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Molecular weight | Value: 0.185 MDa / Experimental value: YES | ||||||||||||||||||||||||||||||
Source (natural) | Organism: Porcine epidemic diarrhea virus | ||||||||||||||||||||||||||||||
Source (recombinant) | Organism: Spodoptera frugiperda (fall armyworm) | ||||||||||||||||||||||||||||||
Buffer solution | pH: 7.5 | ||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3 | ||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-Electron microscopy imaging
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm |
Image recording | Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 103940 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||
Atomic model building |
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Refine LS restraints |
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