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- PDB-7ksg: SARS-CoV-2 spike in complex with nanobodies E -

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Basic information

Entry
Database: PDB / ID: 7ksg
TitleSARS-CoV-2 spike in complex with nanobodies E
Components
  • Nanobody against SARS-CoV-2 glycoprotein
  • Spike glycoproteinPeplomer
KeywordsVIRAL PROTEIN/IMMUNE SYSTEM / SARS-CoV-2 / spike protein / nanobody / VIRUS / VIRAL PROTEIN-IMMUNE SYSTEM complex
Function / homology
Function and homology information


suppression by virus of host tetherin activity / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / receptor-mediated virion attachment to host cell / host cell surface receptor binding / endocytosis involved in viral entry into host cell / fusion of virus membrane with host plasma membrane / suppression by virus of host type I interferon-mediated signaling pathway / viral entry into host cell / fusion of virus membrane with host endosome membrane / viral envelope ...suppression by virus of host tetherin activity / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / receptor-mediated virion attachment to host cell / host cell surface receptor binding / endocytosis involved in viral entry into host cell / fusion of virus membrane with host plasma membrane / suppression by virus of host type I interferon-mediated signaling pathway / viral entry into host cell / fusion of virus membrane with host endosome membrane / viral envelope / pathogenesis / host cell plasma membrane / virion membrane / integral component of membrane / identical protein binding
Coronavirus spike glycoprotein S1, C-terminal / Spike glycoprotein S2 superfamily, coronavirus / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like / Spike glycoprotein, heptad repeat 2, coronavirus / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S1, C-terminal / Spike receptor binding domain superfamily, coronavirus / Betacoronavirus-like spike glycoprotein S1, N-terminal / Betacoronavirus spike glycoprotein S1, receptor binding
Spike glycoprotein
Biological speciesSevere acute respiratory syndrome coronavirus 2
Camelus bactrianus (Bactrian camel)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.33 Å
AuthorsHallberg, B.M. / Das, H.
CitationJournal: Science / Year: 2021
Title: Structure-guided multivalent nanobodies block SARS-CoV-2 infection and suppress mutational escape.
Authors: Paul-Albert Koenig / Hrishikesh Das / Hejun Liu / Beate M Kümmerer / Florian N Gohr / Lea-Marie Jenster / Lisa D J Schiffelers / Yonas M Tesfamariam / Miki Uchima / Jennifer D Wuerth / Karl ...Authors: Paul-Albert Koenig / Hrishikesh Das / Hejun Liu / Beate M Kümmerer / Florian N Gohr / Lea-Marie Jenster / Lisa D J Schiffelers / Yonas M Tesfamariam / Miki Uchima / Jennifer D Wuerth / Karl Gatterdam / Natalia Ruetalo / Maria H Christensen / Caroline I Fandrey / Sabine Normann / Jan M P Tödtmann / Steffen Pritzl / Leo Hanke / Jannik Boos / Meng Yuan / Xueyong Zhu / Jonathan L Schmid-Burgk / Hiroki Kato / Michael Schindler / Ian A Wilson / Matthias Geyer / Kerstin U Ludwig / B Martin Hällberg / Nicholas C Wu / Florian I Schmidt /
Abstract: The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to spread, with devastating consequences. For passive immunization efforts, nanobodies have size and cost ...The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to spread, with devastating consequences. For passive immunization efforts, nanobodies have size and cost advantages over conventional antibodies. In this study, we generated four neutralizing nanobodies that target the receptor binding domain of the SARS-CoV-2 spike protein. We used x-ray crystallography and cryo-electron microscopy to define two distinct binding epitopes. On the basis of these structures, we engineered multivalent nanobodies with more than 100 times the neutralizing activity of monovalent nanobodies. Biparatopic nanobody fusions suppressed the emergence of escape mutants. Several nanobody constructs neutralized through receptor binding competition, whereas other monovalent and biparatopic nanobodies triggered aberrant activation of the spike fusion machinery. These premature conformational changes in the spike protein forestalled productive fusion and rendered the virions noninfectious.
Validation Report
SummaryFull reportAbout validation report
History
DepositionNov 22, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 20, 2021Provider: repository / Type: Initial release

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Structure visualization

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Assembly

Deposited unit
A: Spike glycoprotein
B: Spike glycoprotein
C: Spike glycoprotein
D: Nanobody against SARS-CoV-2 glycoprotein
E: Nanobody against SARS-CoV-2 glycoprotein
F: Nanobody against SARS-CoV-2 glycoprotein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)485,30157
Polymers474,0196
Non-polymers11,28251
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area40170 Å2
ΔGint6 kcal/mol
Surface area165320 Å2

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Components

#1: Protein Spike glycoprotein / Peplomer / S glycoprotein / E2 / Peplomer protein / SARS-CoV-2 spike glycoprotein


Mass: 142399.375 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Severe acute respiratory syndrome coronavirus 2
Gene: S, 2 / Production host: Homo sapiens (human) / References: UniProt: P0DTC2
#2: Antibody Nanobody against SARS-CoV-2 glycoprotein


Mass: 15607.102 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Camelus bactrianus (Bactrian camel) / Production host: Escherichia coli (E. coli)
#3: Sugar...
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 51 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1Complex between SARS-CoV-2 and a neutralising nanobody ECOMPLEX#1-#20RECOMBINANT
2SARS-CoV-2 spike glycoproteinCOMPLEX#11RECOMBINANT
3nanobody ECOMPLEX#21RECOMBINANT
Molecular weightValue: 0.51 MDa / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Severe acute respiratory syndrome coronavirus 22697049
23Camelus bactrianus (Bactrian camel)9837
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Escherichia coli (E. coli)562
Buffer solutionpH: 7.3
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 165000 X / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 1.5 sec. / Electron dose: 49 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1

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Processing

EM software
IDNameVersionCategory
1Warpparticle selection
2EPUimage acquisition
4WarpCTF correction
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
12cryoSPARCclassification
13cryoSPARC2.153D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.33 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 43933 / Symmetry type: POINT

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