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- EMDB-2424: Sub-tomogram average of the glycoprotein spike of Boid Inclusion ... -

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Basic information

Entry
Database: EMDB / ID: EMD-2424
TitleSub-tomogram average of the glycoprotein spike of Boid Inclusion Body Disease associated ArenaVirus
Map dataSub-tomogram reconstruction of the glycoprotein spike from Boid Inclusion Body Disease-associated arenavirus
Sample
  • Sample: Boid inclusion body disease associated arenavirus
  • Protein or peptide: glycoprotein C
KeywordsGlycoprotein Trimer
Biological speciesArenavirus
Methodsubtomogram averaging / cryo EM / Resolution: 32.0 Å
AuthorsHetzel U / Sironen T / Laurinmaki P / Liljeroos L / Patjas A / Henttonen H / Vaheri A / Artelt A / Kipar A / Butcher SJ ...Hetzel U / Sironen T / Laurinmaki P / Liljeroos L / Patjas A / Henttonen H / Vaheri A / Artelt A / Kipar A / Butcher SJ / Vapalahti O / Hepojoki J
CitationJournal: J Virol / Year: 2013
Title: Isolation, identification, and characterization of novel arenaviruses, the etiological agents of boid inclusion body disease.
Authors: Udo Hetzel / Tarja Sironen / Pasi Laurinmäki / Lassi Liljeroos / Aino Patjas / Heikki Henttonen / Antti Vaheri / Annette Artelt / Anja Kipar / Sarah J Butcher / Olli Vapalahti / Jussi Hepojoki /
Abstract: Boid inclusion body disease (BIBD) is a progressive, usually fatal disease of constrictor snakes, characterized by cytoplasmic inclusion bodies (IB) in a wide range of cell types. To identify the ...Boid inclusion body disease (BIBD) is a progressive, usually fatal disease of constrictor snakes, characterized by cytoplasmic inclusion bodies (IB) in a wide range of cell types. To identify the causative agent of the disease, we established cell cultures from BIBD-positive and -negative boa constrictors. The IB phenotype was maintained in cultured cells of affected animals, and supernatants from these cultures caused the phenotype in cultures originating from BIBD-negative snakes. Viruses were purified from the supernatants by ultracentrifugation and subsequently identified as arenaviruses. Purified virus also induced the IB phenotype in naive cells, which fulfilled Koch's postulates in vitro. One isolate, tentatively designated University of Helsinki virus (UHV), was studied in depth. Sequencing confirmed that UHV is a novel arenavirus species that is distinct from other known arenaviruses including those recently identified in snakes with BIBD. The morphology of UHV was established by cryoelectron tomography and subtomographic averaging, revealing the trimeric arenavirus spike structure at 3.2-nm resolution. Immunofluorescence, immunohistochemistry, and immunoblotting with a polyclonal rabbit antiserum against UHV and reverse transcription-PCR (RT-PCR) revealed the presence of genetically diverse arenaviruses in a large cohort of snakes with BIBD, confirming the causative role of arenaviruses. Some snakes were also found to carry arenavirus antibodies. Furthermore, mammalian cells (Vero E6) were productively infected with UHV, demonstrating the potential of arenaviruses to cross species barriers. In conclusion, we propose the newly identified lineage of arenaviruses associated with BIBD as a novel taxonomic entity, boid inclusion body disease-associated arenaviruses (BIBDAV), in the family Arenaviridae.
History
DepositionJul 22, 2013-
Header (metadata) releaseAug 28, 2013-
Map releaseAug 28, 2013-
UpdateOct 9, 2013-
Current statusOct 9, 2013Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.2
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_2424.tif

Downloads & links

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Map

FileDownload / File: emd_2424.map.gz / Format: CCP4 / Size: 1001 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSub-tomogram reconstruction of the glycoprotein spike from Boid Inclusion Body Disease-associated arenavirus
Voxel sizeX=Y=Z: 7.6 Å
Density
Contour LevelBy AUTHOR: 0.2 / Movie #1: 0.2
Minimum - Maximum-229.364013669999991 - 282.838928220000014
Average (Standard dev.)-28.35873604 (±27.214328770000002)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions646464
Spacing646464
CellA=B=C: 486.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z7.67.67.6
M x/y/z646464
origin x/y/z0.0000.0000.000
length x/y/z486.400486.400486.400
α/β/γ90.00090.00090.000
start NX/NY/NZ00-40
NX/NY/NZ555581
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS646464
D min/max/mean-229.364282.839-28.359

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Supplemental data

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Sample components

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Entire : Boid inclusion body disease associated arenavirus

EntireName: Boid inclusion body disease associated arenavirus
Components
  • Sample: Boid inclusion body disease associated arenavirus
  • Protein or peptide: glycoprotein C

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Supramolecule #1000: Boid inclusion body disease associated arenavirus

SupramoleculeName: Boid inclusion body disease associated arenavirus / type: sample / ID: 1000 / Number unique components: 1

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Macromolecule #1: glycoprotein C

MacromoleculeName: glycoprotein C / type: protein_or_peptide / ID: 1 / Name.synonym: GPC
Details: The GPC monomer has a predicted weight of 45 after cleavage of the signal sequence. In most arenaviruses, GPC is cleaved into GP1 (24.1 kDa) and GP2 (20.9 kDa).
Oligomeric state: Trimer / Recombinant expression: No
Source (natural)Organism: Arenavirus / Strain: University of Helsinki virus 1
Molecular weightTheoretical: 45 KDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4 / Details: 0.2M phosphate-buffered saline
GridDetails: C-flat 2/2-4C holey carbon copper grid, glow discharged in air
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER / Method: Blot for 3 seconds before plunging

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 39400 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 6.0 µm / Nominal defocus min: 3.0 µm / Nominal magnification: 39400
Sample stageSpecimen holder: Eucentric / Specimen holder model: GATAN LIQUID NITROGEN / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 °
DateOct 28, 2012
Image recordingCategory: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Details: Data collected with serialEM
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 32.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMOD, PEET, Bsoft
Details: Final maps were calculated from viruses present in 11 different tilt series. Data were split into two halves at the start of the project and refined separately with independent models. The ...Details: Final maps were calculated from viruses present in 11 different tilt series. Data were split into two halves at the start of the project and refined separately with independent models. The FSC was calculated from the comparison of these two independent models.
Number subtomograms used: 2463
DetailsSubtomograms were manually selected from the tomograms by visual inspection

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