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- PDB-5i08: Prefusion structure of a human coronavirus spike protein -

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Basic information

Entry
Database: PDB / ID: 5i08
TitlePrefusion structure of a human coronavirus spike protein
ComponentsSpike glycoprotein,Foldon chimera
KeywordsVIRAL PROTEIN / coronavirus / glycoprotein / prefusion
Function / homology
Function and homology information


: / virion component / endocytosis involved in viral entry into host cell / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / receptor-mediated virion attachment to host cell / membrane => GO:0016020 / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / host cell plasma membrane ...: / virion component / endocytosis involved in viral entry into host cell / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / receptor-mediated virion attachment to host cell / membrane => GO:0016020 / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / host cell plasma membrane / virion membrane / identical protein binding
Similarity search - Function
Spike (S) protein S1 subunit, receptor-binding domain, HKU1-like / : / Spike (S) protein S1 subunit, N-terminal domain, murine hepatitis virus-like / Fibritin C-terminal / Fibritin C-terminal region / Spike glycoprotein S2, coronavirus, C-terminal / Coronavirus spike glycoprotein S2, intravirion / 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 (S) protein S1 subunit, receptor-binding domain, HKU1-like / : / Spike (S) protein S1 subunit, N-terminal domain, murine hepatitis virus-like / Fibritin C-terminal / Fibritin C-terminal region / Spike glycoprotein S2, coronavirus, C-terminal / Coronavirus spike glycoprotein S2, intravirion / 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 / 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
Fibritin / Spike glycoprotein
Similarity search - Component
Biological speciesHuman coronavirus HKU1
Enterobacteria phage T4 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.04 Å
AuthorsKirchdoerfer, R.N. / Cottrell, C.A. / Wang, N. / Pallesen, J. / Yassine, H.M. / Turner, H.L. / Corbett, K.S. / Graham, B.S. / McLellan, J.S. / Ward, A.B.
CitationJournal: Nature / Year: 2016
Title: Pre-fusion structure of a human coronavirus spike protein.
Authors: Robert N Kirchdoerfer / Christopher A Cottrell / Nianshuang Wang / Jesper Pallesen / Hadi M Yassine / Hannah L Turner / Kizzmekia S Corbett / Barney S Graham / Jason S McLellan / Andrew B Ward /
Abstract: HKU1 is a human betacoronavirus that causes mild yet prevalent respiratory disease, and is related to the zoonotic SARS and MERS betacoronaviruses, which have high fatality rates and pandemic ...HKU1 is a human betacoronavirus that causes mild yet prevalent respiratory disease, and is related to the zoonotic SARS and MERS betacoronaviruses, which have high fatality rates and pandemic potential. Cell tropism and host range is determined in part by the coronavirus spike (S) protein, which binds cellular receptors and mediates membrane fusion. As the largest known class I fusion protein, its size and extensive glycosylation have hindered structural studies of the full ectodomain, thus preventing a molecular understanding of its function and limiting development of effective interventions. Here we present the 4.0 Å resolution structure of the trimeric HKU1 S protein determined using single-particle cryo-electron microscopy. In the pre-fusion conformation, the receptor-binding subunits, S1, rest above the fusion-mediating subunits, S2, preventing their conformational rearrangement. Surprisingly, the S1 C-terminal domains are interdigitated and form extensive quaternary interactions that occlude surfaces known in other coronaviruses to bind protein receptors. These features, along with the location of the two protease sites known to be important for coronavirus entry, provide a structural basis to support a model of membrane fusion mediated by progressive S protein destabilization through receptor binding and proteolytic cleavage. These studies should also serve as a foundation for the structure-based design of betacoronavirus vaccine immunogens.
History
DepositionFeb 3, 2016Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 2, 2016Provider: repository / Type: Initial release
Revision 1.1Mar 9, 2016Group: Database references
Revision 1.2Apr 13, 2016Group: Database references
Revision 1.3Dec 18, 2019Group: Other / Category: atom_sites / cell
Item: _atom_sites.fract_transf_matrix[1][1] / _atom_sites.fract_transf_matrix[2][2] ..._atom_sites.fract_transf_matrix[1][1] / _atom_sites.fract_transf_matrix[2][2] / _atom_sites.fract_transf_matrix[3][3] / _cell.Z_PDB
Revision 1.4Apr 22, 2020Group: Database references / Source and taxonomy / Structure summary
Category: entity / entity_name_com ...entity / entity_name_com / entity_src_gen / struct_ref / struct_ref_seq / struct_ref_seq_dif
Item: _entity.pdbx_description / _entity_name_com.name ..._entity.pdbx_description / _entity_name_com.name / _entity_src_gen.pdbx_gene_src_gene / _entity_src_gen.pdbx_gene_src_ncbi_taxonomy_id / _entity_src_gen.pdbx_gene_src_scientific_name / _struct_ref.db_code / _struct_ref.pdbx_align_begin / _struct_ref.pdbx_db_accession / _struct_ref.pdbx_seq_one_letter_code / _struct_ref_seq.db_align_beg / _struct_ref_seq.db_align_end / _struct_ref_seq.pdbx_auth_seq_align_end / _struct_ref_seq.pdbx_db_accession / _struct_ref_seq.seq_align_end

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Structure visualization

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Assembly

Deposited unit
A: Spike glycoprotein,Foldon chimera
B: Spike glycoprotein,Foldon chimera
C: Spike glycoprotein,Foldon chimera


Theoretical massNumber of molelcules
Total (without water)432,8883
Polymers432,8883
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area22100 Å2
ΔGint-56 kcal/mol
Surface area133900 Å2

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Components

#1: Protein Spike glycoprotein,Foldon chimera / HKU1 prefusion spike protein / S glycoprotein / E2 / Peplomer protein / Fibritin / Collar protein / ...HKU1 prefusion spike protein / S glycoprotein / E2 / Peplomer protein / Fibritin / Collar protein / Whisker antigen control protein


Mass: 144295.891 Da / Num. of mol.: 3
Mutation: R751G, R752G, K753S, R754G, R755S,R751G, R752G, K753S, R754G, R755S
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human coronavirus HKU1 (isolate N5), (gene. exp.) Enterobacteria phage T4 (virus)
Strain: isolate N5 / Gene: S, 3, wac / Plasmid: pVRC-8400 / Cell line (production host): FreeStyle 293F / Production host: Homo sapiens (human) / References: UniProt: Q0ZME7, UniProt: P10104

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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: HKU1 spike with attached foldon domain and mutated furin-cleavage site
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.42 MDa / Experimental value: NO
Source (natural)Organism: Human coronavirus HKU1 (isolate N5)
Source (recombinant)Organism: Homo sapiens (human) / Cell: FreeStyle 293F / Plasmid: pVRC8400
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
1200 mMsodium chlorideNaClSodium chloride1
22 mMTrisC4H11NO31
SpecimenConc.: 0.27 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
EM stainingType: NEGATIVE
Details: 3 uL sample was applied to grid for 30 seconds and then blotted. Grids were stained with 3 uL 1% uranyl formate for 60 seconds followed by blotting.
Material: uranyl formate
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: EMS CF-2/2-4C C-Flat
VitrificationInstrument: HOMEMADE PLUNGER / Cryogen name: ETHANE
Details: 3 uL sample was applied to grid, blotted, and plunged into liquid 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 / Nominal magnification: 22500 X / Calibrated magnification: 22500 X / Nominal defocus max: 3500 nm / Nominal defocus min: 1000 nm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 10 sec. / Electron dose: 57 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 1049
Details: Images were collected using Legionon and processed using Appion.
Image scansMovie frames/image: 50

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Processing

EM software
IDNameVersionCategoryDetails
1DoG Pickerparticle selection
2FindEMparticle selection
3Leginonimage acquisition
5CTFFIND3CTF correction
8MODELLERmodel fitting
9Rosetta2015.19model fitting
10Coot0.8.2model fitting
12RELION1.3initial Euler assignment
13RELION1.4b1final Euler assignment
14IMAGICclassificationMSA
15RELION1.4b13D reconstructionrefine
16Rosetta2015.19model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 39164
Details: 2188 particles were selected from a subset of the data using DoG Picker. These particles were used to generate a 3D model from which back projections were derived. Back projection images ...Details: 2188 particles were selected from a subset of the data using DoG Picker. These particles were used to generate a 3D model from which back projections were derived. Back projection images were used as templates for picking particles from the entire dataset using FindEM.
SymmetryPoint symmetry: C3 (3 fold cyclic)
3D reconstructionResolution: 4.04 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 31435 / Algorithm: BACK PROJECTION / Symmetry type: POINT
Atomic model buildingB value: 117 / Protocol: AB INITIO MODEL / Space: REAL / Target criteria: EMRinger
Details: Model building and refinement were conducted using a combination of software programs.

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