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- EMDB-3749: Poliovirus type 3 (strain Saukett) stabilized virus-like particle... -

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Basic information

Entry
Database: EMDB / ID: EMD-3749
TitlePoliovirus type 3 (strain Saukett) stabilized virus-like particle in complex with the pocket factor compound GPP3
Map dataMRC format map file. Contour level determined by visual inspection in UCSF Chimera.
Sample
  • Virus: Human poliovirus 3
    • Protein or peptide: Capsid proteins, VP1
    • Protein or peptide: Capsid proteins, VP2
    • Protein or peptide: Capsid proteins, VP3
    • Protein or peptide: Capsid proteins, VP4
  • Ligand: 1-[5-[4-(ethoxyiminomethyl)phenoxy]-3-methyl-pentyl]-3-pyridin-4-yl-imidazol-2-one
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 / structural molecule activity / virion attachment to host cell
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: 4.1 Å
AuthorsBahar MW / Kotecha A / Fry EE / Stuart DI
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
WHO/Gates foundationRG.IMCB.I8-TSA-083 United Kingdom
CitationJournal: Nat Commun / Year: 2017
Title: Plant-made polio type 3 stabilized VLPs-a candidate synthetic polio vaccine.
Authors: Johanna Marsian / Helen Fox / Mohammad W Bahar / Abhay Kotecha / Elizabeth E Fry / David I Stuart / Andrew J Macadam / David J Rowlands / George P Lomonossoff /
Abstract: Poliovirus (PV) is the causative agent of poliomyelitis, a crippling human disease known since antiquity. PV occurs in two distinct antigenic forms, D and C, of which only the D form elicits a robust ...Poliovirus (PV) is the causative agent of poliomyelitis, a crippling human disease known since antiquity. PV occurs in two distinct antigenic forms, D and C, of which only the D form elicits a robust neutralizing response. Developing a synthetically produced stabilized virus-like particle (sVLP)-based vaccine with D antigenicity, without the drawbacks of current vaccines, will be a major step towards the final eradication of poliovirus. Such a sVLP would retain the native antigenic conformation and the repetitive structure of the original virus particle, but lack infectious genomic material. In this study, we report the production of synthetically stabilized PV VLPs in plants. Mice carrying the gene for the human PV receptor are protected from wild-type PV when immunized with the plant-made PV sVLPs. Structural analysis of the stabilized mutant at 3.6 Å resolution by cryo-electron microscopy and single-particle reconstruction reveals a structure almost indistinguishable from wild-type PV3.Despite the success of current vaccination against poliomyelitis, safe, cheap and effective vaccines remain sought for continuing eradication effort. Here the authors use plants to express stabilized virus-like particles of type 3 poliovirus that can induce a protective immune response in mice transgenic for the human poliovirus receptor.
History
DepositionJun 2, 2017-
Header (metadata) releaseJul 12, 2017-
Map releaseJul 12, 2017-
UpdateSep 21, 2022-
Current statusSep 21, 2022Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0475
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.0475
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-5o5p
  • Surface level: 0.0475
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-5o5p
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3749.png

Downloads & links

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Map

FileDownload / File: emd_3749.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMRC format map file. Contour level determined by visual inspection in UCSF Chimera.
Voxel sizeX=Y=Z: 1.35 Å
Density
Contour LevelBy AUTHOR: 0.0475 / Movie #1: 0.0475
Minimum - Maximum-0.08870073 - 0.14913394
Average (Standard dev.)0.00028353283 (±0.006716631)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 691.2 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.351.351.35
M x/y/z512512512
origin x/y/z0.0000.0000.000
length x/y/z691.200691.200691.200
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS512512512
D min/max/mean-0.0890.1490.000

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Supplemental data

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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 proteins, VP1
    • Protein or peptide: Capsid proteins, VP2
    • Protein or peptide: Capsid proteins, VP3
    • Protein or peptide: Capsid proteins, VP4
  • Ligand: 1-[5-[4-(ethoxyiminomethyl)phenoxy]-3-methyl-pentyl]-3-pyridin-4-yl-imidazol-2-one

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Supramolecule #1: Human poliovirus 3

SupramoleculeName: Human poliovirus 3 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Details: Poliovirus type 3 (Saukett strain) virus-like particle produced in plant expression system.
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: Human poliovirus 3 / Strain: Saukett
Host systemOrganism: Nicotiana benthamiana (plant) / Recombinant plasmid: pEAQ-HT
Molecular weightTheoretical: 5.8 MDa
Virus shellShell ID: 1 / Name: Capsid / Diameter: 327.0 Å / T number (triangulation number): 1

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Macromolecule #1: Capsid proteins, VP1

MacromoleculeName: Capsid proteins, VP1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human poliovirus 3 / Strain: Saukett
Molecular weightTheoretical: 33.562785 KDa
Recombinant expressionOrganism: Nicotiana benthamiana (plant)
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

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Macromolecule #2: Capsid proteins, VP2

MacromoleculeName: Capsid proteins, VP2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human poliovirus 3 / Strain: Saukett
Molecular weightTheoretical: 30.188982 KDa
Recombinant expressionOrganism: Nicotiana benthamiana (plant)
SequenceString: SPNVEACGYS DRVLQLTIGN STITTQEAAN SVVAYGRWPE FIRDDEANPV DQPTEPDVAT CRFYTLDTVM WGKESKGWWW KLPDALRDM GLFGQNMYYH YLGRSGYTVH VQCNASKFHQ GALGVFAIPE YCLAGDSDKQ RYTSYANANP GEKGGKFYSQ F NRDTAVTS ...String:
SPNVEACGYS DRVLQLTIGN STITTQEAAN SVVAYGRWPE FIRDDEANPV DQPTEPDVAT CRFYTLDTVM WGKESKGWWW KLPDALRDM GLFGQNMYYH YLGRSGYTVH VQCNASKFHQ GALGVFAIPE YCLAGDSDKQ RYTSYANANP GEKGGKFYSQ F NRDTAVTS PKREFCPVDY LLGCGVLLGN AFVYPHQIIN LRTNNSATIV LPYVNAMAID SMVKHNNWGI AILPLSPLDF AQ ESSVEIP ITVTIAPMCS EFNGLRNVTA PKFQ

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Macromolecule #3: Capsid proteins, VP3

MacromoleculeName: Capsid proteins, VP3 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human poliovirus 3 / Strain: Saukett
Molecular weightTheoretical: 26.3151 KDa
Recombinant expressionOrganism: Nicotiana benthamiana (plant)
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

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Macromolecule #4: Capsid proteins, VP4

MacromoleculeName: Capsid proteins, VP4 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human poliovirus 3 / Strain: Saukett
Molecular weightTheoretical: 7.452113 KDa
Recombinant expressionOrganism: Nicotiana benthamiana (plant)
SequenceString:
MGAQVSSQKV GAHENSNRAY GGSTINYTTI NYYKDSASNA ASKQDYSQDP SKFTEPLKDV LIKTAPALN

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Macromolecule #5: 1-[5-[4-(ethoxyiminomethyl)phenoxy]-3-methyl-pentyl]-3-pyridin-4-...

MacromoleculeName: 1-[5-[4-(ethoxyiminomethyl)phenoxy]-3-methyl-pentyl]-3-pyridin-4-yl-imidazol-2-one
type: ligand / ID: 5 / Number of copies: 1 / Formula: 9LW
Molecular weightTheoretical: 408.493 Da
Chemical component information

ChemComp-9LW:
1-[5-[4-(ethoxyiminomethyl)phenoxy]-3-methyl-pentyl]-3-pyridin-4-yl-imidazol-2-one

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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

Concentration0.1 mg/mL
BufferpH: 7 / Details: PBS pH 7.0
GridModel: C-flat CF-2/1-2C / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK I / Details: Blot for 4 seconds before plunging..
DetailsVirus-like particles for polio type 3 (strain Saukett) in complex with pocket factor compound GPP3 were assessed by negative stain EM analysis, prior to cryo-em data collection.

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Electron microscopy

MicroscopeFEI POLARA 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated magnification: 37037 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 2.8000000000000003 µm / Nominal defocus min: 0.8 µm
Specialist opticsEnergy filter - Name: GIF Quantum / Energy filter - Lower energy threshold: 0 eV / Energy filter - Upper energy threshold: 20 eV
Sample stageSpecimen holder model: GATAN 910 MULTI-SPECIMEN SINGLE TILT CRYO TRANSFER HOLDER
Cooling holder cryogen: NITROGEN
DetailsPreliminary grid screening was performed.
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4086 pixel / Digitization - Dimensions - Height: 4086 pixel / Digitization - Sampling interval: 5.0 µm / Digitization - Frames/image: 2-25 / Number grids imaged: 1 / Number real images: 1018 / Average exposure time: 0.2 sec. / Average electron dose: 1.2 e/Å2
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 6639
Details: Automated particle picking (ETHAN) was followed by manual cleaning (EMAN2).
CTF correctionSoftware - Name: RELION (ver. 1.3) / Software - details: CTFFIND3 module within RELION
Details: CTF parameters were estimated using CTFFIND3 as part of RELION 1.3.
Startup modelType of model: EMDB MAP
EMDB ID:

3747


Details: Low pass filtered to 40 Angstroms.
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 1.3)
Final 3D classificationNumber classes: 4 / Software - Name: RELION (ver. 1.3) / Details: 2060 particles selected after 3D classification.
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 1.3)
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.3) / Number images used: 2060
DetailsThe selected images were drift corrected using MotionCorr.
FSC plot (resolution estimation)

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Atomic model buiding 1

Detailsphenix.real_space_refine was used to refine the atomic model in the cryo-em map.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation coefficient
Output model

PDB-5o5p:
Poliovirus type 3 (strain Saukett) stabilized virus-like particle in complex with the pocket factor compound GPP3

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