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- EMDB-36904: SARS-CoV-2 spike protein in complex with a trivalent nanobody -

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Basic information

Entry
Database: EMDB / ID: EMD-36904
TitleSARS-CoV-2 spike protein in complex with a trivalent nanobody
Map dataSARS-CoV-2 spike protein in complex with a trivalant nanobody
Sample
  • Complex: SARS-CoV-2 spike protein in complex with a trivalent nanobody
    • Complex: Trivalent nanobody
    • Complex: SARS-CoV-2 spike protein
Keywordstrivalent nanobody / viral neutralization / all-RBD-down spike / 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 / receptor-mediated endocytosis of virus by host cell / membrane fusion / Attachment and Entry / positive regulation of viral entry into host cell / receptor-mediated virion attachment to host cell / receptor ligand activity / symbiont-mediated suppression of host innate immune response / host cell surface receptor binding / 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 / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike glycoprotein, betacoronavirus / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Betacoronavirus spike glycoprotein S1, receptor binding / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like ...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. / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike glycoprotein, betacoronavirus / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Betacoronavirus spike glycoprotein S1, receptor binding / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like / Betacoronavirus-like spike glycoprotein S1, N-terminal / Spike glycoprotein S2, coronavirus, heptad repeat 1 / Spike glycoprotein S2, coronavirus, heptad repeat 2 / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 2 (HR2) region profile. / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 1 (HR1) region profile. / Spike glycoprotein S2 superfamily, coronavirus / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S2
Similarity search - Domain/homology
Biological speciesSevere acute respiratory syndrome coronavirus 2 / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsJiang XY / Qian JQ / Zhu HX / Qin Q / Huang Q
Funding support China, 2 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31971377 China
National Natural Science Foundation of China (NSFC)31671386 China
CitationJournal: Int J Biol Macromol / Year: 2024
Title: Structure-guided design of a trivalent nanobody cluster targeting SARS-CoV-2 spike protein.
Authors: Xinyi Jiang / Qin Qin / Haixia Zhu / Jiaqiang Qian / Qiang Huang /
Abstract: Nanobodies are natural anti-SARS-CoV-2 drug candidates. Engineering multivalent nanobodies is an effective way to improve the functional binding affinity of natural nanobodies by simultaneously ...Nanobodies are natural anti-SARS-CoV-2 drug candidates. Engineering multivalent nanobodies is an effective way to improve the functional binding affinity of natural nanobodies by simultaneously targeting multiple sites on viral proteins. However, multivalent nanobodies have usually been engineered by trial and error, and rational designs are still lacking. Here, we describe a structure-guided design of a self-assembled trivalent nanobody cluster targeting the SARS-CoV-2 spike protein. Using the nanobody Nb6 as a monovalent binder, we first selected a human-derived trimerization scaffold evaluated by molecular dynamics simulations, then selected an optimal linker according to the minimum distance between Nb6 and the trimerization scaffold, and finally successfully engineered a trivalent nanobody cluster called Tribody. Compared with the low-affinity monovalent counterpart (Nb6), Tribody showed much higher target binding affinity (K < 1 pM) and thus had a 900-fold increase in antiviral neutralization against SARS-CoV-2 pseudovirus. We determined the cryo-EM structure of the Tribody-spike complex and confirmed that all three Nb6 binders of Tribody collectively bind to the three receptor-binding domains (RBDs) of the spike and lock them in a 3-RBD-down conformation, fully consistent with our structure-guided design. This study demonstrates that synthetic nanobody clusters with human-derived self-assembled scaffolds are potential protein drugs against SARS-CoV-2 coronaviruses.
History
DepositionJul 21, 2023-
Header (metadata) releaseJul 24, 2024-
Map releaseJul 24, 2024-
UpdateJul 24, 2024-
Current statusJul 24, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_36904.png

Downloads & links

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Map

FileDownload / File: emd_36904.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSARS-CoV-2 spike protein in complex with a trivalant nanobody
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.57 Å/pix.
x 256 pix.
= 401.92 Å		1.57 Å/pix.
x 256 pix.
= 401.92 Å		1.57 Å/pix.
x 256 pix.
= 401.92 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.57 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.0
Minimum - Maximum-0.5058891 - 2.1747677
Average (Standard dev.)-0.0064510275 (±0.1159665)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 401.92 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_36904_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Sharpen Map

Fileemd_36904_additional_1.map
AnnotationSharpen Map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B

Fileemd_36904_half_map_1.map
AnnotationHalf map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A

Fileemd_36904_half_map_2.map
AnnotationHalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : SARS-CoV-2 spike protein in complex with a trivalent nanobody

EntireName: SARS-CoV-2 spike protein in complex with a trivalent nanobody
Components
  • Complex: SARS-CoV-2 spike protein in complex with a trivalent nanobody
    • Complex: Trivalent nanobody
    • Complex: SARS-CoV-2 spike protein

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Supramolecule #1: SARS-CoV-2 spike protein in complex with a trivalent nanobody

SupramoleculeName: SARS-CoV-2 spike protein in complex with a trivalent nanobody
type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2

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Supramolecule #2: Trivalent nanobody

SupramoleculeName: Trivalent nanobody / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: synthetic construct (others)

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Supramolecule #3: SARS-CoV-2 spike protein

SupramoleculeName: SARS-CoV-2 spike protein / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation state3D array

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Sample preparation

Concentration0.3 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClsodium chloride
20.0 mMC4H11NO3tris(hydroxymethyl) aminomethane

Details: 20 mM Tris,150 mM NaCl
GridModel: Quantifoil / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 296.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS GLACIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 56.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.8 µm / Nominal magnification: 92000

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Image processing

Particle selectionNumber selected: 2754775
Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.1.2) / Number images used: 262855
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.1.2)
Final angle assignmentType: OTHER / Software - Name: cryoSPARC (ver. 4.1.2)
FSC plot (resolution estimation)

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