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- PDB-8ydy: Cryo-EM structure of SARS-CoV-2 spike ectodomain (HexaPro, Omicro... -

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Basic information

Entry
Database: PDB / ID: 8ydy
TitleCryo-EM structure of SARS-CoV-2 spike ectodomain (HexaPro, Omicron BA.5 variant) in complex with CeSPIACE, class 1
Components
  • CeSPIACE
  • Spike glycoprotein
KeywordsVIRAL PROTEIN/INHIBITOR / Spike protein / VIRAL PROTEIN-INHIBITOR COMPLEX
Function / homology
Function and homology information


Maturation of spike protein / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / viral translation / symbiont-mediated-mediated suppression of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / membrane fusion ...Maturation of spike protein / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / viral translation / symbiont-mediated-mediated suppression of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / membrane fusion / entry receptor-mediated virion attachment to host cell / Attachment and Entry / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / positive regulation of viral entry into host cell / receptor-mediated virion attachment to host cell / host cell surface receptor binding / symbiont-mediated suppression of host innate immune response / receptor ligand activity / endocytosis involved in viral entry into host cell / 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 / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal / Spike glycoprotein, betacoronavirus / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus ...Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal / Spike glycoprotein, betacoronavirus / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / 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 1 (HR1) region profile. / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 2 (HR2) region profile. / Spike glycoprotein S2 superfamily, coronavirus / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S2
Similarity search - Domain/homology
Biological speciesSevere acute respiratory syndrome coronavirus 2
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 8 Å
AuthorsSuzuki, H. / Nakamura, S. / Fujiyoshi, Y.
Funding support Japan, 5items
OrganizationGrant numberCountry
Japan Agency for Medical Research and Development (AMED)JP21fk0108462 Japan
Japan Agency for Medical Research and Development (AMED)JP21fk0108585 Japan
Japan Agency for Medical Research and Development (AMED)JP21ae0121028 Japan
Japan Agency for Medical Research and Development (AMED)JP18ae0101046 Japan
Japan Society for the Promotion of Science (JSPS)20H00451 Japan
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Structure-guided engineering of a mutation-tolerant inhibitor peptide against variable SARS-CoV-2 spikes.
Authors: Shun Nakamura / Yukihiro Tanimura / Risa Nomura / Hiroshi Suzuki / Kouki Nishikawa / Akiko Kamegawa / Nobutaka Numoto / Atsushi Tanaka / Shigeru Kawabata / Shoichi Sakaguchi / Akino Emi / ...Authors: Shun Nakamura / Yukihiro Tanimura / Risa Nomura / Hiroshi Suzuki / Kouki Nishikawa / Akiko Kamegawa / Nobutaka Numoto / Atsushi Tanaka / Shigeru Kawabata / Shoichi Sakaguchi / Akino Emi / Youichi Suzuki / Yoshinori Fujiyoshi /
Abstract: Pathogen mutations present an inevitable and challenging problem for therapeutics and the development of mutation-tolerant anti-infective drugs to strengthen global health and combat evolving ...Pathogen mutations present an inevitable and challenging problem for therapeutics and the development of mutation-tolerant anti-infective drugs to strengthen global health and combat evolving pathogens is urgently needed. While spike proteins on viral surfaces are attractive targets for preventing viral entry, they mutate frequently, making it difficult to develop effective therapeutics. Here, we used a structure-guided strategy to engineer an inhibitor peptide against the SARS-CoV-2 spike, called CeSPIACE, with mutation-tolerant and potent binding ability against all variants to enhance affinity for the invariant architecture of the receptor-binding domain (RBD). High-resolution structures of the peptide complexed with mutant RBDs revealed a mechanism of mutation-tolerant inhibition. CeSPIACE bound major mutant RBDs with picomolar affinity and inhibited infection by SARS-CoV-2 variants in VeroE6/TMPRSS2 cells (IC 4 pM to 13 nM) and demonstrated potent in vivo efficacy by inhalation administration in hamsters. Mutagenesis analyses to address mutation risks confirmed tolerance against existing and/or potential future mutations of the RBD. Our strategy of engineering mutation-tolerant inhibitors may be applicable to other infectious diseases.
History
DepositionFeb 21, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 15, 2025Provider: repository / Type: Initial release
Revision 1.1Jul 30, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Spike glycoprotein
B: Spike glycoprotein
C: Spike glycoprotein
D: CeSPIACE
E: CeSPIACE
F: CeSPIACE
hetero molecules


Theoretical massNumber of molelcules
Total (without water)447,63010
Polymers446,7456
Non-polymers8854
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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


Mass: 144193.672 Da / Num. of mol.: 3 / Mutation: F817P, A892P, A899P, A942P, K986P, V987P
Source method: isolated from a genetically manipulated source
Details: the N-terminal sequence (from -18 to 12) is an mammalian secretion signal sequence, and the C-terminal sequence includes "T4 fibritin trimerization mottif" (Gly1211-Leu1237), "HRV 3C ...Details: the N-terminal sequence (from -18 to 12) is an mammalian secretion signal sequence, and the C-terminal sequence includes "T4 fibritin trimerization mottif" (Gly1211-Leu1237), "HRV 3C protease cleavage site" (Leu1241-Pro1248), "8x His tag" (His1250-His1257) and "Twin-strep tag" (Trp1260-Lys1288)
Source: (gene. exp.) Severe acute respiratory syndrome coronavirus 2
Gene: S, 2 / Cell line (production host): Expi293F / Production host: Homo sapiens (human) / References: UniProt: P0DTC2
#2: Protein/peptide CeSPIACE


Mass: 4721.475 Da / Num. of mol.: 3 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#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


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 4 / 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
Has protein modificationY

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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: SARS-CoV-2 spike ectodomain (HexaPro, Omicron BA.5 variant) in complex with CeSPIACE
Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
Buffer component
IDConc.NameBuffer-ID
12 mMTris-HCl1
2200 mMsodium chloride1
30.02 %sodium azide1
SpecimenConc.: 0.42 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 278 K

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Electron microscopy imaging

MicroscopyModel: JEOL CRYO ARM 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm / Cs: 3.4 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: JEOL 3200FSC CRYOHOLDER
Image recordingElectron dose: 69.6 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM software
IDNameVersionCategory
1Topazparticle selection
2Gautomatchparticle selection
3SerialEM3.8.9image acquisition
5CTFFINDCTF correction
8UCSF Chimeramodel fitting
9UCSF ChimeraXmodel fitting
11Cootmodel refinement
12RELION4initial Euler assignment
13RELION4final Euler assignment
15RELION43D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 18473 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT
Atomic model buildingPDB-ID: 6XKL

6xkl


Accession code: 6XKL / Source name: PDB / Type: experimental model

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