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- EMDB-22139: SARS-CoV-2 ORF3a with Emodin in a MSP1E3D1 lipid nanodisc -

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Basic information

Entry
Database: EMDB / ID: EMD-22139
TitleSARS-CoV-2 ORF3a with Emodin in a MSP1E3D1 lipid nanodisc
Map data
SampleORF3a in MSPE3D1 nanodisc:
nucleic-acidNucleic acid
Biological speciesSevere acute respiratory syndrome coronavirus 2
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsKern DM / Hoel CM / Brohawn SG
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM128263 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM123496 United States
CitationJournal: bioRxiv / Year: 2020
Title: Cryo-EM structure of the SARS-CoV-2 3a ion channel in lipid nanodiscs.
Authors: David M Kern / Ben Sorum / Christopher M Hoel / Savitha Sridharan / Jonathan P Remis / Daniel B Toso / Stephen G Brohawn
Abstract: SARS-CoV-2 encodes three putative ion channels: E, 8a, and 3a. In related SARS-CoV-1, 3a is implicated in viral release, inflammasome activation, and cell death and its deletion reduces viral titer ...SARS-CoV-2 encodes three putative ion channels: E, 8a, and 3a. In related SARS-CoV-1, 3a is implicated in viral release, inflammasome activation, and cell death and its deletion reduces viral titer and morbidity in animal models, suggesting 3a-targeted therapeutics could treat SARS and COVID-19. However, the structural basis for the function of 3a is unknown. Here, we show that SARS-CoV-2 forms large conductance cation channels and present cryo-EM structures of dimeric and tetrameric SARS-CoV-2 3a in lipid nanodiscs. 3a adopts a novel fold and is captured in a closed or inactivated state. A narrow bifurcated exterior pore precludes conduction and leads to a large polar cavity open to the cytosol. 3a function is conserved in a common variant among circulating SARS-CoV-2 that alters the channel pore. We identify 3a-like proteins in and that infect bats and humans, suggesting therapeutics targeting 3a could treat a range of coronaviral diseases.
History
DepositionJun 11, 2020-
Header (metadata) releaseJun 17, 2020-
Map releaseJun 17, 2020-
UpdateJun 17, 2020-
Current statusJun 17, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.845
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.845
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_22139.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.14 Å/pix.
x 256 pix.
= 291.072 Å
1.14 Å/pix.
x 256 pix.
= 291.072 Å
1.14 Å/pix.
x 256 pix.
= 291.072 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.137 Å
Density
Contour LevelBy AUTHOR: 0.845 / Movie #1: 0.845
Minimum - Maximum-3.0434036 - 4.0708475
Average (Standard dev.)-0.00047682092 (±0.06503829)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 291.072 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.1371.1371.137
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z291.072291.072291.072
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ360360360
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-3.0434.071-0.000

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Supplemental data

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Sample components

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Entire ORF3a in MSPE3D1 nanodisc

EntireName: ORF3a in MSPE3D1 nanodisc / Number of components: 2

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Component #1: protein, ORF3a in MSPE3D1 nanodisc

ProteinName: ORF3a in MSPE3D1 nanodisc / Recombinant expression: No
MassTheoretical: 64 kDa
SourceSpecies: Severe acute respiratory syndrome coronavirus 2
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #2: nucleic-acid, SARS-CoV-2 ORF3a

nucleic acidName: SARS-CoV-2 ORF3a / Class: OTHER / Structure: OTHER / Synthetic: No
Sequence: MDLFMRIFTI GTVTLKQGEI KDATPSDFVR ATATIPIQAS LPFGWLIVGV ALLAVFQSAS KIITLKKRWQ LALSKGVHFV CNLLLLFVTV YSHLLLVAAG LEAPFLYLYA LVYFLQSINF VRIIMRLWLC WKCRSKNPLL YDANYFLCWH TNCYDYCIPY NSVTSSIVIT ...Sequence:
MDLFMRIFTI GTVTLKQGEI KDATPSDFVR ATATIPIQAS LPFGWLIVGV ALLAVFQSAS KIITLKKRWQ LALSKGVHFV CNLLLLFVTV YSHLLLVAAG LEAPFLYLYA LVYFLQSINF VRIIMRLWLC WKCRSKNPLL YDANYFLCWH TNCYDYCIPY NSVTSSIVIT SGDGTTSPIS EHDYQIGGYT EKWESGVKDC VVLHSYFTSD YYQLYSTQLS TDTGVEHVTF FIYNKIVDEP EEHVQIHTID GSSGVVNPVM EPIYDEPTTT TSVPLSNSLE VLFQ
SourceSpecies: Severe acute respiratory syndrome coronavirus 2

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 1.1 mg/mL / pH: 7.4
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277 K / Humidity: 100 %
Details: 1 blot force, 5 second wait time, 3 second blot time.

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
ImagingMicroscope: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 47.20275 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: OTHER

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Image acquisition

Image acquisitionNumber of digital images: 6750 / Details: 50 frames per image

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 51908
3D reconstructionSoftware: cryoSPARC / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Target criteria: Correlation coefficient / Refinement space: REAL
Input PDB model: 6XDC
Overall bvalue: 212.6

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