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- EMDB-11330: Structure of Disulphide-stabilized SARS-CoV-2 Spike Protein Trime... -

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Entry
Database: EMDB / ID: EMD-11330
TitleStructure of Disulphide-stabilized SARS-CoV-2 Spike Protein Trimer (x1 disulphide-bond mutant, S383C, D985C, K986P, V987P, single Arg S1/S2 cleavage site) in Closed State
Map dataLocal Resolution Filtered Sharpened Map
Sample
  • Complex: Disulphide-stabilized SARS-CoV-2 Spike protein trimer
    • Protein or peptide: Spike glycoprotein
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
Keywordscoronavirus / SARS-CoV-2 / Spike protein / S protein / S antigen / COVID-19 / receptor binding / membrane fusion / vaccine design / VIRAL PROTEIN
Function / homology
Function and homology information


Maturation of spike protein / viral translation / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / suppression by virus of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / entry receptor-mediated virion attachment to host cell ...Maturation of spike protein / viral translation / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / suppression by virus of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / entry receptor-mediated virion attachment to host cell / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / membrane fusion / receptor-mediated endocytosis of virus by host cell / Attachment and Entry / positive regulation of viral entry into host cell / receptor-mediated virion attachment to host cell / receptor ligand activity / host cell surface receptor binding / symbiont-mediated suppression of host innate immune response / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / SARS-CoV-2 activates/modulates innate and adaptive immune responses / host cell plasma membrane / virion membrane / identical protein binding / membrane / plasma membrane
Similarity search - Function
Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal / Spike glycoprotein, betacoronavirus / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus ...Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal / Spike glycoprotein, betacoronavirus / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like / Betacoronavirus-like spike glycoprotein S1, N-terminal / Betacoronavirus spike glycoprotein S1, receptor binding / Spike glycoprotein S2, coronavirus, heptad repeat 1 / Spike glycoprotein S2, coronavirus, heptad repeat 2 / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 1 (HR1) region profile. / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 2 (HR2) region profile. / Spike glycoprotein S2 superfamily, coronavirus / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S2
Similarity search - Domain/homology
Biological speciesSevere acute respiratory syndrome coronavirus 2
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsQu K / Xiong X / Scheres SHW / Briggs JAG
Funding support United Kingdom, European Union, Germany, 4 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MC_UP_1201/16 United Kingdom
European Research Council (ERC)ERC-CoG-648432European Union
German Research Foundation (DFG)240245660 - SFB1129 Germany
Medical Research Council (MRC, United Kingdom)MC_UP_A025_1013 United Kingdom
CitationJournal: Nat Struct Mol Biol / Year: 2020
Title: A thermostable, closed SARS-CoV-2 spike protein trimer.
Authors: Xiaoli Xiong / Kun Qu / Katarzyna A Ciazynska / Myra Hosmillo / Andrew P Carter / Soraya Ebrahimi / Zunlong Ke / Sjors H W Scheres / Laura Bergamaschi / Guinevere L Grice / Ying Zhang / / ...Authors: Xiaoli Xiong / Kun Qu / Katarzyna A Ciazynska / Myra Hosmillo / Andrew P Carter / Soraya Ebrahimi / Zunlong Ke / Sjors H W Scheres / Laura Bergamaschi / Guinevere L Grice / Ying Zhang / / James A Nathan / Stephen Baker / Leo C James / Helen E Baxendale / Ian Goodfellow / Rainer Doffinger / John A G Briggs /
Abstract: The spike (S) protein of SARS-CoV-2 mediates receptor binding and cell entry and is the dominant target of the immune system. It exhibits substantial conformational flexibility. It transitions from ...The spike (S) protein of SARS-CoV-2 mediates receptor binding and cell entry and is the dominant target of the immune system. It exhibits substantial conformational flexibility. It transitions from closed to open conformations to expose its receptor-binding site and, subsequently, from prefusion to postfusion conformations to mediate fusion of viral and cellular membranes. S-protein derivatives are components of vaccine candidates and diagnostic assays, as well as tools for research into the biology and immunology of SARS-CoV-2. Here we have designed mutations in S that allow the production of thermostable, disulfide-bonded S-protein trimers that are trapped in the closed, prefusion state. Structures of the disulfide-stabilized and non-disulfide-stabilized proteins reveal distinct closed and locked conformations of the S trimer. We demonstrate that the designed, thermostable, closed S trimer can be used in serological assays. This protein has potential applications as a reagent for serology, virology and as an immunogen.
History
DepositionJul 8, 2020-
Header (metadata) releaseJul 22, 2020-
Map releaseJul 22, 2020-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0211
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.0211
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6zoy
  • Surface level: 0.0211
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_11330.png

Downloads & links

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Map

FileDownload / File: emd_11330.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLocal Resolution Filtered Sharpened Map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 256 pix.
= 271.616 Å		1.06 Å/pix.
x 256 pix.
= 271.616 Å		1.06 Å/pix.
x 256 pix.
= 271.616 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.061 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0211 / Movie #1: 0.0211
Minimum - Maximum-0.09588575 - 0.19248168
Average (Standard dev.)0.00021608135 (±0.0052638724)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 271.616 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0611.0611.061
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z271.616271.616271.616
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ250250250
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0960.1920.000

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Supplemental data

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Additional map: Unsharpened Map

Fileemd_11330_additional.map
AnnotationUnsharpened Map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Disulphide-stabilized SARS-CoV-2 Spike protein trimer

EntireName: Disulphide-stabilized SARS-CoV-2 Spike protein trimer
Components
  • Complex: Disulphide-stabilized SARS-CoV-2 Spike protein trimer
    • Protein or peptide: Spike glycoprotein
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: Disulphide-stabilized SARS-CoV-2 Spike protein trimer

SupramoleculeName: Disulphide-stabilized SARS-CoV-2 Spike protein trimer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: S383C, D985C, K986P, V987P, Single Arg S1/S2 cleavage site
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2 / Strain: WIV-04
Molecular weightTheoretical: 28 KDa

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Macromolecule #1: Spike glycoprotein

MacromoleculeName: Spike glycoprotein / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2
Molecular weightTheoretical: 138.282938 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: ETGTQCVNLT TRTQLPPAYT NSFTRGVYYP DKVFRSSVLH STQDLFLPFF SNVTWFHAIH VSGTNGTKRF DNPVLPFNDG VYFASTEKS NIIRGWIFGT TLDSKTQSLL IVNNATNVVI KVCEFQFCND PFLGVYYHKN NKSWMESEFR VYSSANNCTF E YVSQPFLM ...String:
ETGTQCVNLT TRTQLPPAYT NSFTRGVYYP DKVFRSSVLH STQDLFLPFF SNVTWFHAIH VSGTNGTKRF DNPVLPFNDG VYFASTEKS NIIRGWIFGT TLDSKTQSLL IVNNATNVVI KVCEFQFCND PFLGVYYHKN NKSWMESEFR VYSSANNCTF E YVSQPFLM DLEGKQGNFK NLREFVFKNI DGYFKIYSKH TPINLVRDLP QGFSALEPLV DLPIGINITR FQTLLALHRS YL TPGDSSS GWTAGAAAYY VGYLQPRTFL LKYNENGTIT DAVDCALDPL SETKCTLKSF TVEKGIYQTS NFRVQPTESI VRF PNITNL CPFGEVFNAT RFASVYAWNR KRISNCVADY SVLYNSASFS TFKCYGVCPT KLNDLCFTNV YADSFVIRGD EVRQ IAPGQ TGKIADYNYK LPDDFTGCVI AWNSNNLDSK VGGNYNYLYR LFRKSNLKPF ERDISTEIYQ AGSTPCNGVE GFNCY FPLQ SYGFQPTNGV GYQPYRVVVL SFELLHAPAT VCGPKKSTNL VKNKCVNFNF NGLTGTGVLT ESNKKFLPFQ QFGRDI ADT TDAVRDPQTL EILDITPCSF GGVSVITPGT NTSNQVAVLY QDVNCTEVPV AIHADQLTPT WRVYSTGSNV FQTRAGC LI GAEHVNNSYE CDIPIGAGIC ASYQTQTNSR SVASQSIIAY TMSLGAENSV AYSNNSIAIP TNFTISVTTE ILPVSMTK T SVDCTMYICG DSTECSNLLL QYGSFCTQLN RALTGIAVEQ DKNTQEVFAQ VKQIYKTPPI KDFGGFNFSQ ILPDPSKPS KRSFIEDLLF NKVTLADAGF IKQYGDCLGD IAARDLICAQ KFNGLTVLPP LLTDEMIAQY TSALLAGTIT SGWTFGAGAA LQIPFAMQM AYRFNGIGVT QNVLYENQKL IANQFNSAIG KIQDSLSSTA SALGKLQDVV NQNAQALNTL VKQLSSNFGA I SSVLNDIL SRLCPPEAEV QIDRLITGRL QSLQTYVTQQ LIRAAEIRAS ANLAATKMSE CVLGQSKRVD FCGKGYHLMS FP QSAPHGV VFLHVTYVPA QEKNFTTAPA ICHDGKAHFP REGVFVSNGT HWFVTQRNFY EPQIITTDNT FVSGNCDVVI GIV NNTVYD PLQPELDSFK EELDKYFKNH TSPDVDLGDI SGINASVVNI QKEIDRLNEV AKNLNESLID LQELGKYEQY IKGS GRENL YFQGGGGSGY IPEAPRDGQA YVRKDGEWVL LSTFLGHHHH HH

UniProtKB: Spike glycoprotein

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Macromolecule #3: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 3 / Number of copies: 36 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.69 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
148.0 mMNaClsodium chloride
8.0 mMNa2HPO4disodium hydrogen phospahte
2.0 mMKH2PO4potassium dihydrogen phospahte
0.01 %C14H28O6octyl glucoside
GridModel: C-flat-2/2 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 2441 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 81000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 587351 / Details: template picking
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 210946
Initial angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 3.1)
Final 3D classificationNumber classes: 8 / Software - Name: RELION (ver. 3.1)

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Atomic model buiding 1

Initial modelPDB ID:

6vxx


Chain - Chain ID: A / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 116 / Target criteria: correlation coefficient
Output model

PDB-6zoy:
Structure of Disulphide-stabilized SARS-CoV-2 Spike Protein Trimer (x1 disulphide-bond mutant, S383C, D985C, K986P, V987P, single Arg S1/S2 cleavage site) in Closed State

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