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- PDB-8ajl: Structure of the Ancestral Scaffold Antigen-6 of Coronavirus Spik... -

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Basic information

Entry
Database: PDB / ID: 8ajl
TitleStructure of the Ancestral Scaffold Antigen-6 of Coronavirus Spike protein
ComponentsSpike glycoprotein,Fibritin
KeywordsBIOSYNTHETIC PROTEIN / Ancestor / Spike / Coronavirus / Scaffold / S-protein / Protein Engineering
Function / homology
Function and homology information


virion component / 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 ...virion component / 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
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. / 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 ...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. / 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
Spike glycoprotein / Fibritin
Similarity search - Component
Biological speciesSevere acute respiratory syndrome coronavirus
Enterobacteria phage T4 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.77 Å
AuthorsHueting, D. / Schriever, K. / Wallden, K. / Andrell, J. / Syren, P.O.
Funding support Sweden, European Union, 5items
OrganizationGrant numberCountry
Other government2019-700
Other governmentSNIC 2021/5-70
The Swedish Foundation for Strategic ResearchFFF20-0027 Sweden
European Union (EU)ATAC, 101003650European Union
Knut and Alice Wallenberg FoundationSciLifeLab/KAW national COVID-19 research program project grant 2020 (KAW 2020.0182) Sweden
Citation
Journal: Nat Commun / Year: 2023
Title: Design, structure and plasma binding of ancestral β-CoV scaffold antigens.
Authors: David Hueting / Karen Schriever / Rui Sun / Stelios Vlachiotis / Fanglei Zuo / Likun Du / Helena Persson / Camilla Hofström / Mats Ohlin / Karin Walldén / Marcus Buggert / Lennart ...Authors: David Hueting / Karen Schriever / Rui Sun / Stelios Vlachiotis / Fanglei Zuo / Likun Du / Helena Persson / Camilla Hofström / Mats Ohlin / Karin Walldén / Marcus Buggert / Lennart Hammarström / Harold Marcotte / Qiang Pan-Hammarström / Juni Andréll / Per-Olof Syrén /
Abstract: We report the application of ancestral sequence reconstruction on coronavirus spike protein, resulting in stable and highly soluble ancestral scaffold antigens (AnSAs). The AnSAs interact with plasma ...We report the application of ancestral sequence reconstruction on coronavirus spike protein, resulting in stable and highly soluble ancestral scaffold antigens (AnSAs). The AnSAs interact with plasma of patients recovered from COVID-19 but do not bind to the human angiotensin-converting enzyme 2 (ACE2) receptor. Cryo-EM analysis of the AnSAs yield high resolution structures (2.6-2.8 Å) indicating a closed pre-fusion conformation in which all three receptor-binding domains (RBDs) are facing downwards. The structures reveal an intricate hydrogen-bonding network mediated by well-resolved loops, both within and across monomers, tethering the N-terminal domain and RBD together. We show that AnSA-5 can induce and boost a broad-spectrum immune response against the wild-type RBD as well as circulating variants of concern in an immune organoid model derived from tonsils. Finally, we highlight how AnSAs are potent scaffolds by replacing the ancestral RBD with the wild-type sequence, which restores ACE2 binding and increases the interaction with convalescent plasma.
#1: Journal: Res Sq / Year: 2023
Title: Design, structure and plasma binding of ancestral beta-CoV scaffold antigens
Authors: Hueting, D. / Schriever, K. / Zuo, F. / Du, L. / Persson, H. / Hofstrom, C. / Ohlin, M. / Wallden, K. / Hammarstrom, L. / Marcotte, H. / Pan-Hammarstrom, Q. / Andrell, J. / Syren, P.O.
History
DepositionJul 28, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 16, 2023Provider: repository / Type: Initial release
Revision 1.1Aug 30, 2023Group: Database references / Category: citation
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.year
Revision 1.2Nov 1, 2023Group: Database references / Refinement description / Category: citation / citation_author / em_3d_fitting_list
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Spike glycoprotein,Fibritin
C: Spike glycoprotein,Fibritin
B: Spike glycoprotein,Fibritin
hetero molecules


Theoretical massNumber of molelcules
Total (without water)430,75457
Polymers416,9803
Non-polymers13,77454
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area52830 Å2
ΔGint63 kcal/mol
Surface area134930 Å2
MethodPISA

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Components

#1: Protein Spike glycoprotein,Fibritin / S glycoprotein / E2 / Peplomer protein / Collar protein / Whisker antigen control protein


Mass: 138993.469 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Details: AnSA-6 full sequence 1-19 WT SARS-CoV-2 S protein signaling peptide 672-679 No density. 1139 onwards, No density. Expression tags.,AnSA-6 full sequence 1-19 WT SARS-CoV-2 S protein signaling ...Details: AnSA-6 full sequence 1-19 WT SARS-CoV-2 S protein signaling peptide 672-679 No density. 1139 onwards, No density. Expression tags.,AnSA-6 full sequence 1-19 WT SARS-CoV-2 S protein signaling peptide 672-679 No density. 1139 onwards, No density. Expression tags.
Source: (gene. exp.) Severe acute respiratory syndrome coronavirus, (gene. exp.) Enterobacteria phage T4 (Bacteriophage T4) (virus)
Gene: S, 2, wac / Plasmid: pMx series / Production host: Homo sapiens (human) / Strain (production host): Expi293F / References: UniProt: P0DTC2, UniProt: P10104
#2: 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.: 9
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-Glycam 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
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}}LINUCSPDB-CARE
#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.: 45 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
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 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

ComponentName: Ancestral S-protein of coronaviruses related to SARS-CoV-2
Type: COMPLEX
Details: Ancestral coronavirus generated by Ancestral sequence reconstruction expressed in human cells.
Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Severe acute respiratory syndrome coronavirus2901879
31Enterobacteria phage T4 (Bacteriophage T4) (virus)10665
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDStrain
21Homo sapiens (human)9606Expi293F
31Homo sapiens (human)9606Expi293F
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESHEPES1
2100 mMsodium chlorideNaClSodium chloride1
SpecimenConc.: 1.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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: OTHER
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 105000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 600 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 1.11 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameVersionCategory
1cryoSPARC3.3.1particle selection
2EPUv2.11.1image acquisition
4cryoSPARC3.3.1CTF correction
7ISOLDE1.1model fitting
9PHENIX1.3model refinement
13ISOLDE1.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 2035826
3D reconstructionResolution: 2.77 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 253429 / Symmetry type: POINT
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
16ZOZ

6zoz

16ZOZ1PDBexperimental model
27BNN

7bnn

17BNN2PDBexperimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00627354
ELECTRON MICROSCOPYf_angle_d0.68537248
ELECTRON MICROSCOPYf_dihedral_angle_d12.6449861
ELECTRON MICROSCOPYf_chiral_restr0.0494401
ELECTRON MICROSCOPYf_plane_restr0.0094755

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