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- PDB-7ntc: Trimeric SARS-CoV-2 spike ectodomain bound to P008_056 Fab -

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Basic information

Entry
Database: PDB / ID: 7ntc
TitleTrimeric SARS-CoV-2 spike ectodomain bound to P008_056 Fab
Components
  • (P008_056 Fab ...) x 2
  • Spike glycoproteinSpike protein
KeywordsVIRAL PROTEIN / SARS-CoV-2 / spike / biliverdin / COVID19 / Antibody / Fab / epitope / NTD
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 / Attachment and Entry / membrane fusion / positive regulation of viral entry into host cell / receptor-mediated virion attachment to host cell / receptor ligand activity / host cell surface receptor binding / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / virion attachment to host cell / SARS-CoV-2 activates/modulates innate and adaptive immune responses / host cell plasma membrane / virion membrane / membrane / identical protein binding / 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 / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S2 / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal
Similarity search - Domain/homology
BILIVERDINE IX ALPHA / Spike glycoprotein
Similarity search - Component
Biological speciesSevere acute respiratory syndrome coronavirus 2
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsRosa, A. / Pye, V.E. / Nans, A. / Cherepanov, P.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
The Francis Crick InstituteFC001061 United Kingdom
CitationJournal: Sci Adv / Year: 2021
Title: SARS-CoV-2 can recruit a heme metabolite to evade antibody immunity.
Authors: Annachiara Rosa / Valerie E Pye / Carl Graham / Luke Muir / Jeffrey Seow / Kevin W Ng / Nicola J Cook / Chloe Rees-Spear / Eleanor Parker / Mariana Silva Dos Santos / Carolina Rosadas / ...Authors: Annachiara Rosa / Valerie E Pye / Carl Graham / Luke Muir / Jeffrey Seow / Kevin W Ng / Nicola J Cook / Chloe Rees-Spear / Eleanor Parker / Mariana Silva Dos Santos / Carolina Rosadas / Alberto Susana / Hefin Rhys / Andrea Nans / Laura Masino / Chloe Roustan / Evangelos Christodoulou / Rachel Ulferts / Antoni G Wrobel / Charlotte-Eve Short / Michael Fertleman / Rogier W Sanders / Judith Heaney / Moira Spyer / Svend Kjær / Andy Riddell / Michael H Malim / Rupert Beale / James I MacRae / Graham P Taylor / Eleni Nastouli / Marit J van Gils / Peter B Rosenthal / Massimo Pizzato / Myra O McClure / Richard S Tedder / George Kassiotis / Laura E McCoy / Katie J Doores / Peter Cherepanov /
Abstract: The coronaviral spike is the dominant viral antigen and the target of neutralizing antibodies. We show that SARS-CoV-2 spike binds biliverdin and bilirubin, the tetrapyrrole products of heme ...The coronaviral spike is the dominant viral antigen and the target of neutralizing antibodies. We show that SARS-CoV-2 spike binds biliverdin and bilirubin, the tetrapyrrole products of heme metabolism, with nanomolar affinity. Using cryo-electron microscopy and x-ray crystallography, we mapped the tetrapyrrole interaction pocket to a deep cleft on the spike N-terminal domain (NTD). At physiological concentrations, biliverdin significantly dampened the reactivity of SARS-CoV-2 spike with immune sera and inhibited a subset of neutralizing antibodies. Access to the tetrapyrrole-sensitive epitope is gated by a flexible loop on the distal face of the NTD. Accompanied by profound conformational changes in the NTD, antibody binding requires relocation of the gating loop, which folds into the cleft vacated by the metabolite. Our results indicate that SARS-CoV-2 spike NTD harbors a dominant epitope, access to which can be controlled by an allosteric mechanism that is regulated through recruitment of a metabolite.
History
DepositionMar 9, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 28, 2021Provider: repository / Type: Initial release
Revision 1.1May 5, 2021Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.2Jun 9, 2021Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.title ..._citation.journal_volume / _citation.title / _citation_author.identifier_ORCID / _citation_author.name

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Structure visualization

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  • Deposited structure unit
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  • Superimposition on EM map
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Structure viewerMolecule:
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Assembly

Deposited unit
A: Spike glycoprotein
B: Spike glycoprotein
C: Spike glycoprotein
H: P008_056 Fab Heavy chain
L: P008_056 Fab Light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)497,49361
Polymers479,3025
Non-polymers18,19056
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: microscopy, by cryoEM
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area49990 Å2
ΔGint47 kcal/mol
Surface area180170 Å2
MethodPISA

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Components

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Antibody , 2 types, 2 molecules HL

#2: Antibody P008_056 Fab Heavy chain


Mass: 27239.400 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell (production host): Expi293 / Production host: Homo sapiens (human)
#3: Antibody P008_056 Fab Light chain


Mass: 25253.180 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell (production host): Expi293 / Production host: Homo sapiens (human)

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Protein / Non-polymers , 2 types, 5 molecules ABC

#1: Protein Spike glycoprotein / Spike protein / S glycoprotein / E2 / Peplomer protein


Mass: 142269.859 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 / Cell (production host): Expi293 / Production host: Homo sapiens (human) / References: UniProt: P0DTC2
#5: Chemical ChemComp-BLA / BILIVERDINE IX ALPHA


Mass: 582.646 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C33H34N4O6 / Feature type: SUBJECT OF INVESTIGATION

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Sugars , 2 types, 54 molecules

#4: Polysaccharide...
2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 25 / Source method: obtained synthetically
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#6: 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.: 29 / 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

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Details

Has ligand of interestY

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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
IDNameTypeDetailsEntity IDParent-IDSource
1Stabilised SARS-CoV-2 spike ectodomain trimer bound to one P008_056 FabCOMPLEXnatural source below: Spike is SARS-Cov-2 and Fab is Human - there is only option for one natural source below!#1-#30MULTIPLE SOURCES
2Stabilised SARS-CoV-2 spike ectodomain trimerCOMPLEX#11RECOMBINANT
3P008_056 FabCOMPLEX#2-#31RECOMBINANT
Molecular weightValue: 0.47886 MDa / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Severe acute respiratory syndrome coronavirus 22697049
33Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDCell
22Homo sapiens (human)9606Expi293
33Homo sapiens (human)9606Expi293
Buffer solutionpH: 8
Details: 150 mM NaCl, 1 mM ethylenediaminetetraacetic acid (EDTA), 25 mM Tris-HCl, pH 8.0
SpecimenConc.: 0.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: SARS-CoV-2 spike ectodomain (0.6 mg/ml final concentration in TBSE supplemented with 0.1% n-octylglucoside) with 0.2 mg/ml P008_056 Fab
Specimen supportGrid type: Quantifoil R2/2
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 51 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 17010
Details: A total of 17010 movies were collected with a pixel size of 1.38 Angstrom and total electron exposure of 51 electrons per square Angstrom, spread over 40 frames.

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Processing

SoftwareName: PHENIX / Version: 1.19rc5_4047: / Classification: refinement
EM software
IDNameVersionCategory
7UCSF Chimeramodel fitting
12cryoSPARC23D reconstruction
13PHENIX1.19rc5-4047model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 78291 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL
Details: in addition to the spike: 6APC and 6PHB were used as templates for the heavy and light chains of the Fab respectively
Atomic model buildingPDB-ID: 6ZGE

6zge

Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00430686
ELECTRON MICROSCOPYf_angle_d0.55241770
ELECTRON MICROSCOPYf_dihedral_angle_d6.64531
ELECTRON MICROSCOPYf_chiral_restr0.0464924
ELECTRON MICROSCOPYf_plane_restr0.0045365

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