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- EMDB-15543: Poliovirus type 3 (strain Saukett) stabilised virus-like particle... -

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Basic information

Entry
Database: EMDB / ID: EMD-15543
TitlePoliovirus type 3 (strain Saukett) stabilised virus-like particle (PV3 SC8).
Map dataHalf map after RELION 3D refinement. Contour level recommended based on viewing in UCSF chimeraX
Sample
  • Virus: Human poliovirus 3
    • Protein or peptide: Capsid protein, VP1
    • Protein or peptide: Capsid protein, VP0
    • Protein or peptide: Capsid protein, VP3
  • Ligand: SPHINGOSINE
KeywordsCapsid protein / VIRUS LIKE PARTICLE
Function / homology
Function and homology information


symbiont genome entry into host cell via pore formation in plasma membrane / viral capsid / symbiont-mediated suppression of host gene expression / virion attachment to host cell / structural molecule activity
Similarity search - Function
Picornavirus coat protein VP4 / Picornavirus coat protein (VP4) / Picornavirus capsid / picornavirus capsid protein / Picornavirus/Calicivirus coat protein / Viral coat protein subunit
Similarity search - Domain/homology
Biological speciesHuman poliovirus 3
Methodsingle particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsBahar MW / Fry EE / Stuart DI
Funding support United States, 1 items
OrganizationGrant numberCountry
Bill & Melinda Gates FoundationRG.IMCB.I8-TSA-083 United States
CitationJournal: Viruses / Year: 2022
Title: Production and Characterisation of Stabilised PV-3 Virus-like Particles Using .
Authors: Lee Sherry / Keith Grehan / Jessica J Swanson / Mohammad W Bahar / Claudine Porta / Elizabeth E Fry / David I Stuart / David J Rowlands / Nicola J Stonehouse /
Abstract: Following the success of global vaccination programmes using the live-attenuated oral and inactivated poliovirus vaccines (OPV and IPV), wild poliovirus (PV) is now only endemic in Afghanistan and ...Following the success of global vaccination programmes using the live-attenuated oral and inactivated poliovirus vaccines (OPV and IPV), wild poliovirus (PV) is now only endemic in Afghanistan and Pakistan. However, the continued use of these vaccines poses potential risks to the eradication of PV. The production of recombinant PV virus-like particles (VLPs), which lack the viral genome offer great potential as next-generation vaccines for the post-polio world. We have previously reported production of PV VLPs using , however, these VLPs were in the non-native conformation (C Ag), which would not produce effective protection against PV. Here, we build on this work and show that it is possible to produce wt PV-3 and thermally stabilised PV-3 (referred to as PV-3 SC8) VLPs in the native conformation (D Ag) using . We show that the PV-3 SC8 VLPs provide a much-improved D:C antigen ratio as compared to wt PV-3, whilst exhibiting greater thermostability than the current IPV vaccine. Finally, we determine the cryo-EM structure of the yeast-derived PV-3 SC8 VLPs and compare this to previously published PV-3 D Ag structures, highlighting the similarities between these recombinantly expressed VLPs and the infectious virus, further emphasising their potential as a next-generation vaccine candidate for PV.
History
DepositionAug 6, 2022-
Header (metadata) releaseOct 12, 2022-
Map releaseOct 12, 2022-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15543.png

Downloads & links

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Map

FileDownload / File: emd_15543.map.gz / Format: CCP4 / Size: 347.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationHalf map after RELION 3D refinement. Contour level recommended based on viewing in UCSF chimeraX
Voxel sizeX=Y=Z: 0.82 Å
Density
Contour LevelBy AUTHOR: 0.06
Minimum - Maximum-0.13615467 - 0.23435937
Average (Standard dev.)0.0017614996 (±0.015325013)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions450450450
Spacing450450450
CellA=B=C: 369.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half map after RELION 3D refinement. Contour level...

Fileemd_15543_half_map_1.map
AnnotationHalf map after RELION 3D refinement. Contour level recommended based on viewing in UCSF chimeraX.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map after RELION 3D refinement. Contour level...

Fileemd_15543_half_map_2.map
AnnotationHalf map after RELION 3D refinement. Contour level recommended based on viewing in UCSF chimeraX.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Human poliovirus 3

EntireName: Human poliovirus 3
Components
  • Virus: Human poliovirus 3
    • Protein or peptide: Capsid protein, VP1
    • Protein or peptide: Capsid protein, VP0
    • Protein or peptide: Capsid protein, VP3
  • Ligand: SPHINGOSINE

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Supramolecule #1: Human poliovirus 3

SupramoleculeName: Human poliovirus 3 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Details: Recombinantly expressed virus-like particle of PV3 (strain Saukett).
NCBI-ID: 12086 / Sci species name: Human poliovirus 3 / Sci species strain: Saukett / Virus type: VIRUS-LIKE PARTICLE / Virus isolate: SEROTYPE / Virus enveloped: No / Virus empty: Yes
Host (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 5.85 MDa
Virus shellShell ID: 1 / Name: Virus shell 1 / Diameter: 310.0 Å / T number (triangulation number): 1

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Macromolecule #1: Capsid protein, VP1

MacromoleculeName: Capsid protein, VP1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human poliovirus 3
Molecular weightTheoretical: 33.562785 KDa
Recombinant expressionOrganism: Komagataella pastoris (fungus)
SequenceString: GIEDLITEVA QGALTLSLPK QQDSLPDTKA SGPAHSKEVP ALTAVETGAT NPLVPSDTVQ TRHVIQRRSR SESTIESFFA RGACVAIIE VDNEEPTTRA QKLFAMWRIT YKDTVQLRRK LEFFTYSRFD MELTFVVTAN FTNTNNGHAL NQVYQIMYIP P GAPTPKSW ...String:
GIEDLITEVA QGALTLSLPK QQDSLPDTKA SGPAHSKEVP ALTAVETGAT NPLVPSDTVQ TRHVIQRRSR SESTIESFFA RGACVAIIE VDNEEPTTRA QKLFAMWRIT YKDTVQLRRK LEFFTYSRFD MELTFVVTAN FTNTNNGHAL NQVYQIMYIP P GAPTPKSW DDYTWQTSSN PSIFYTYGAA PARISVPYVG LANAYSHFYD GFAKVPLKTD ANDQIGDSLY SAMTVDDFGV LA IRVVNDH NPTKVTSKVR IYMKPKHVRV WCPRPPRAVP YYGPGVDYKD NLNPLSEKGL TTY

UniProtKB: Genome polyprotein

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Macromolecule #2: Capsid protein, VP0

MacromoleculeName: Capsid protein, VP0 / type: protein_or_peptide / ID: 2
Details: Sequence is given for the VP0 polypeptide. Mutations are numbered according to sequence numbering for mature polypeptides VP2 and VP4.
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human poliovirus 3
Molecular weightTheoretical: 37.623023 KDa
Recombinant expressionOrganism: Komagataella pastoris (fungus)
SequenceString: MGAQVSSQKV GAHENSNRAY GGSTINYTTI NYYKDSASNA ASKQDYSQDP SKFTEPLKDV LIKTAPALNS PNVEACGYSD RVLQLTIGN STITTQEAAN SVVAYGRWPE FIRDDEANPV DQPTEPDVAT CRFYTLDTVM WGKESKGWWW KLPDALRDMG L FGQNMYYH ...String:
MGAQVSSQKV GAHENSNRAY GGSTINYTTI NYYKDSASNA ASKQDYSQDP SKFTEPLKDV LIKTAPALNS PNVEACGYSD RVLQLTIGN STITTQEAAN SVVAYGRWPE FIRDDEANPV DQPTEPDVAT CRFYTLDTVM WGKESKGWWW KLPDALRDMG L FGQNMYYH YLGRSGYTVH VQCNASKFHQ GALGVFAIPE YCLAGDSDKQ RYTSYANANP GEKGGKFYSQ FNRDTAVTSP KR EFCPVDY LLGCGVLLGN AFVYPHQIIN LRTNNSATIV LPYVNAMAID SMVKHNNWGI AILPLSPLDF AQESSVEIPI TVT IAPMCS EFNGLRNVTA PKFQ

UniProtKB: Genome polyprotein

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Macromolecule #3: Capsid protein, VP3

MacromoleculeName: Capsid protein, VP3 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human poliovirus 3
Molecular weightTheoretical: 26.3151 KDa
Recombinant expressionOrganism: Komagataella pastoris (fungus)
SequenceString: GLPVLNTPGS NQYLTSDNYQ SPCAIPEFDV TPPIDIPGEV KNMMELAEID TMIPLNLENT KRNTMDMYRV TLSDSADLSQ PILCFSLSP ASDPRLSHTM LGEVLNYYTH WAGSLKFTFL FCGSMMATGK ILVAYAPPGA QPPTSRKEAM LGTHVIWDLG L QSSCTMVV ...String:
GLPVLNTPGS NQYLTSDNYQ SPCAIPEFDV TPPIDIPGEV KNMMELAEID TMIPLNLENT KRNTMDMYRV TLSDSADLSQ PILCFSLSP ASDPRLSHTM LGEVLNYYTH WAGSLKFTFL FCGSMMATGK ILVAYAPPGA QPPTSRKEAM LGTHVIWDLG L QSSCTMVV PWISNVTYRQ TTQDSFTEGG YISMFYQTRI VVPLSTPKSM SMLGFVSACN DFSVRLLRDT THISQSALPQ

UniProtKB: Genome polyprotein

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Macromolecule #4: SPHINGOSINE

MacromoleculeName: SPHINGOSINE / type: ligand / ID: 4 / Number of copies: 1 / Formula: SPH
Molecular weightTheoretical: 299.492 Da
Chemical component information

ChemComp-SPH:
SPHINGOSINE / Sphingosine

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.5 mg/mL
BufferpH: 7
Component:
ConcentrationName
1.0 xDPBS
20.0 mMEDTAEthylenediaminetetraacetic acid

Details: 1 x DPBS, 20 mM EDTA, pH 7.0
GridModel: EMS Lacey Carbon / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: LACEY / Support film - Film thickness: 3 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR
Details: The specific type of grid used was Ultra-thin carbon support film, 3nm - on lacey carbon AGS187-4 from Agar Scientific.
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
Details: Double blotting with 4 ul of sample, followed by 4 second blot, before plunging..
DetailsVirus-like particle purified by sucrose density gradient purification from Pichia pastoris cells.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated magnification: 60975 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 165000
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-20 / Number grids imaged: 1 / Number real images: 3717 / Average exposure time: 3.0 sec. / Average electron dose: 35.0 e/Å2 / Details: Pixel sampling of 0.82 A/pixel.
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 10744
Startup modelType of model: EMDB MAP
EMDB ID:

3747


Details: Startup model was low pass filtered to 60 Angstroms.
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.1)
Final 3D classificationNumber classes: 5 / Avg.num./class: 829 / Software - Name: RELION (ver. 3.1.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.1)
Final reconstructionNumber classes used: 2 / Applied symmetry - Point group: I (icosahedral) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1.1)
Details: Final reconstruction was sharpened with Post-processing in RELION using an inverse B-factor of -69.1 Angstroms.
Number images used: 2712
DetailsPixels size was 0.82 A/pixel
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6z6w


Chain - Source name: PDB / Chain - Initial model type: experimental model
DetailsInitial model was rigid body fitted using UCSF chimera and Coot. Global minimization and B-factor refinement was performed in real space using phenix_real.space.refine.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Correlation coefficient
Output model

PDB-8anw:
Poliovirus type 3 (strain Saukett) stabilised virus-like particle (PV3 SC8).

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