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- EMDB-11719: Cryo-EM structure of a prefusion stabilized SARS-CoV-2 Spike (D61... -

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Basic information

Entry
Database: EMDB / ID: EMD-11719
TitleCryo-EM structure of a prefusion stabilized SARS-CoV-2 Spike (D614N, R682S, R685G, A892P, A942P and V987P)(One up trimer)
Map data
SampleS closed protein trimer
  • Spike glycoprotein,Envelope glycoprotein,Spike glycoprotein,Envelope glycoprotein,SARS-CoV-2 S protein
  • ligand
Function / homology
Function and homology information


Maturation of spike protein / Translation of Structural Proteins / Virion Assembly and Release / suppression by virus of host tetherin activity / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / Attachment and Entry / receptor-mediated virion attachment to host cell / endoplasmic reticulum-Golgi intermediate compartment / host cell surface receptor binding / endocytosis involved in viral entry into host cell ...Maturation of spike protein / Translation of Structural Proteins / Virion Assembly and Release / suppression by virus of host tetherin activity / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / Attachment and Entry / receptor-mediated virion attachment to host cell / endoplasmic reticulum-Golgi intermediate compartment / host cell surface receptor binding / endocytosis involved in viral entry into host cell / endocytic vesicle membrane / fusion of virus membrane with host plasma membrane / suppression by virus of host type I interferon-mediated signaling pathway / fusion of virus membrane with host endosome membrane / viral entry into host cell / viral envelope / endoplasmic reticulum lumen / host cell plasma membrane / virion membrane / integral component of membrane / identical protein binding
Similarity search - Function
Fibritin C-terminal / Fibritin C-terminal region / Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike glycoprotein, N-terminal domain superfamily / Spike glycoprotein S2, coronavirus, heptad repeat 1 / Spike glycoprotein S2, coronavirus, heptad repeat 2 / Spike glycoprotein, betacoronavirus ...Fibritin C-terminal / Fibritin C-terminal region / Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike glycoprotein, N-terminal domain superfamily / Spike glycoprotein S2, coronavirus, heptad repeat 1 / Spike glycoprotein S2, coronavirus, heptad repeat 2 / Spike glycoprotein, betacoronavirus / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike receptor binding domain superfamily, coronavirus / Betacoronavirus spike glycoprotein S1, receptor binding / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 1 (HR1) region profile. / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like / Betacoronavirus-like spike glycoprotein S1, N-terminal / Spike glycoprotein S2 superfamily, coronavirus / Coronavirus spike glycoprotein S2 / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal
Similarity search - Domain/homology
Envelope glycoprotein / Spike glycoprotein
Similarity search - Component
Biological speciesSevere acute respiratory syndrome coronavirus 2
Methodsingle particle reconstruction / cryo EM / Resolution: 2.92 Å
AuthorsRenault LLR / Rutten L / Juraszek J / Langedijk JPM
CitationJournal: Nat Commun / Year: 2021
Title: Stabilizing the closed SARS-CoV-2 spike trimer.
Authors: Jarek Juraszek / Lucy Rutten / Sven Blokland / Pascale Bouchier / Richard Voorzaat / Tina Ritschel / Mark J G Bakkers / Ludovic L R Renault / Johannes P M Langedijk /
Abstract: The trimeric spike (S) protein of SARS-CoV-2 is the primary focus of most vaccine design and development efforts. Due to intrinsic instability typical of class I fusion proteins, S tends to ...The trimeric spike (S) protein of SARS-CoV-2 is the primary focus of most vaccine design and development efforts. Due to intrinsic instability typical of class I fusion proteins, S tends to prematurely refold to the post-fusion conformation, compromising immunogenic properties and prefusion trimer yields. To support ongoing vaccine development efforts, we report the structure-based design of soluble S trimers with increased yields and stabilities, based on introduction of single point mutations and disulfide-bridges. We identify regions critical for stability: the heptad repeat region 1, the SD1 domain and position 614 in SD2. We combine a minimal selection of mostly interprotomeric mutations to create a stable S-closed variant with a 6.4-fold higher expression than the parental construct while no longer containing a heterologous trimerization domain. The cryo-EM structure reveals a correctly folded, predominantly closed pre-fusion conformation. Highly stable and well producing S protein and the increased understanding of S protein structure will support vaccine development and serological diagnostics.
History
DepositionSep 14, 2020-
Header (metadata) releaseNov 4, 2020-
Map releaseNov 4, 2020-
UpdateJan 27, 2021-
Current statusJan 27, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.012
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.012
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7ad1
  • Surface level: 0.012
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_11719.map.gz / Format: CCP4 / Size: 343 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 448 pix.
= 374.528 Å
0.84 Å/pix.
x 448 pix.
= 374.528 Å
0.84 Å/pix.
x 448 pix.
= 374.528 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.836 Å
Density
Contour LevelBy AUTHOR: 0.009 / Movie #1: 0.012
Minimum - Maximum-0.052745935 - 0.09797727
Average (Standard dev.)0.00011210258 (±0.001627465)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions448448448
Spacing448448448
CellA=B=C: 374.528 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.8360.8360.836
M x/y/z448448448
origin x/y/z0.0000.0000.000
length x/y/z374.528374.528374.528
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ400400400
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS448448448
D min/max/mean-0.0530.0980.000

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Supplemental data

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Sample components

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Entire S closed protein trimer

EntireName: S closed protein trimer / Number of Components: 3

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Component #1: cellular-component, S closed protein trimer

Cellular-componentName: S closed protein trimer / Recombinant expression: No
SourceSpecies: Severe acute respiratory syndrome coronavirus 2
Source (engineered)Expression System: Homo sapiens (human) / Vector: pCDNA2004 / Cell of expression system: kidney

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Component #2: protein, Spike glycoprotein,Envelope glycoprotein,Spike glycoprot...

ProteinName: Spike glycoprotein,Envelope glycoprotein,Spike glycoprotein,Envelope glycoprotein,SARS-CoV-2 S protein
Number of Copies: 3 / Recombinant expression: No
MassTheoretical: 142.547344 kDa
SourceSpecies: Severe acute respiratory syndrome coronavirus 2
Source (engineered)Expression System: Homo sapiens (human)

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Component #3: ligand, 2-acetamido-2-deoxy-beta-D-glucopyranose

LigandName: 2-acetamido-2-deoxy-beta-D-glucopyranose / Number of Copies: 10 / Recombinant expression: No
MassTheoretical: 0.221208 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen State: Particle / Method: cryo EM
Sample solutionpH: 7
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen Name: ETHANE / Temperature: 295 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron Source: FIELD EMISSION GUN / Accelerating Voltage: 300 kV / Electron Dose: 65 e/Å2 / Illumination Mode: OTHER
LensMagnification: 105000 X (nominal), 105000 X (calibrated) / Cs: 2.7 mm / Imaging Mode: BRIGHT FIELD / Defocus: 600.0 - 2500.0 nm / Energy Filter: GIF Quantum LS
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: OTHER

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Image acquisition

Image acquisitionNumber of Digital Images: 9760
Details: Movies with 50 frames were acquired in super resolution counting mode.

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Image processing

ProcessingMethod: single particle reconstruction / Number of Projections: 184000
Details: collected movies were subjected to beam induced drift correction using MotionCor2 and downsampled at this stage
3D reconstructionSoftware: RELION / Resolution: 2.92 Å / Resolution Method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Output model

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