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- EMDB-4092: Tomographic reconstruction of ex vivo mammalian prions -

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Basic information

Entry
Database: EMDB / ID: EMD-4092
TitleTomographic reconstruction of ex vivo mammalian prions
Map dataNone
Sample
  • Organelle or cellular component: Infectious prion rods
Biological speciesMus musculus (house mouse)
Methodelectron tomography / negative staining
AuthorsTerry C / Wenborn A / Gros N / Sells J / Joiner S / Hosszu LLP / Tattum MH / Panico S / Clare DK / Collinge J ...Terry C / Wenborn A / Gros N / Sells J / Joiner S / Hosszu LLP / Tattum MH / Panico S / Clare DK / Collinge J / Saibil HR / Wadsworth JDF
CitationJournal: Open Biol / Year: 2016
Title: Ex vivo mammalian prions are formed of paired double helical prion protein fibrils.
Authors: Cassandra Terry / Adam Wenborn / Nathalie Gros / Jessica Sells / Susan Joiner / Laszlo L P Hosszu / M Howard Tattum / Silvia Panico / Daniel K Clare / John Collinge / Helen R Saibil / Jonathan D F Wadsworth /
Abstract: Mammalian prions are hypothesized to be fibrillar or amyloid forms of prion protein (PrP), but structures observed to date have not been definitively correlated with infectivity and the three- ...Mammalian prions are hypothesized to be fibrillar or amyloid forms of prion protein (PrP), but structures observed to date have not been definitively correlated with infectivity and the three-dimensional structure of infectious prions has remained obscure. Recently, we developed novel methods to obtain exceptionally pure preparations of prions from mouse brain and showed that pathogenic PrP in these high-titre preparations is assembled into rod-like assemblies. Here, we have used precise cell culture-based prion infectivity assays to define the physical relationship between the PrP rods and prion infectivity and have used electron tomography to define their architecture. We show that infectious PrP rods isolated from multiple prion strains have a common hierarchical assembly comprising twisted pairs of short fibres with repeating substructure. The architecture of the PrP rods provides a new structural basis for understanding prion infectivity and can explain the inability to systematically generate high-titre synthetic prions from recombinant PrP.
History
DepositionAug 4, 2016-
Header (metadata) releaseAug 17, 2016-
Map releaseAug 17, 2016-
UpdateJul 12, 2017-
Current statusJul 12, 2017Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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Supplemental images

Downloads & links

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Map

FileDownload / File: emd_4092.map.gz / Format: CCP4 / Size: 23.7 MB / Type: IMAGE STORED AS SIGNED BYTE
AnnotationNone
Voxel sizeX=Y=Z: 4.153 Å
Density
Minimum - Maximum-128. - 127.
Average (Standard dev.)80.924614000000005 (±75.538284000000004)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin0020
Dimensions60060069
Spacing60060069
CellA: 2491.7998 Å / B: 2491.7998 Å / C: 286.557 Å
α=β=γ: 90.0 °

CCP4 map header:

modeenvelope stored as signed bytes (from -128 lowest to 127 highest)
Å/pix. X/Y/Z4.1534.1534.153
M x/y/z60060069
origin x/y/z0.0000.0000.000
length x/y/z2491.8002491.800286.557
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS0020
NC/NR/NS60060069
D min/max/mean-128.000127.00080.925

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Supplemental data

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Sample components

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Entire : Infectious prion rods

EntireName: Infectious prion rods
Components
  • Organelle or cellular component: Infectious prion rods

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Supramolecule #1: Infectious prion rods

SupramoleculeName: Infectious prion rods / type: organelle_or_cellular_component / ID: 1 / Parent: 0
Details: Ex vivo, isolated from prion-infected mouse brain (prion strain ME7)
Source (natural)Organism: Mus musculus (house mouse) / Strain: C57Bl/6 / Organ: Brain

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Experimental details

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Structure determination

Methodnegative staining
Processingelectron tomography
Aggregation statefilament

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Sample preparation

BufferpH: 7.4
Component:
ConcentrationName
20.0 mMTris
150.0 mMSodium chloride
StainingType: NEGATIVE / Material: Uranyl Acetate / Details: 2% (w/v)
GridModel: Electron Microscopy Sciences / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 5.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
DetailsPrions isolated from mouse brain, approximate concentration 3 ug/ml
SectioningOther: NO SECTIONING

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Electron microscopy

MicroscopeFEI TECNAI 12
Electron beamAcceleration voltage: 120 kV / Electron source: TUNGSTEN HAIRPIN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated defocus max: 1.0 µm / Calibrated defocus min: 0.5 µm / Calibrated magnification: 58594 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 1.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 42000
Sample stageSpecimen holder model: OTHER
Image recordingFilm or detector model: GATAN MULTISCAN / Digitization - Dimensions - Width: 1024 pixel / Digitization - Dimensions - Height: 1024 pixel / Digitization - Sampling interval: 15.0 µm / Number grids imaged: 1 / Number real images: 1 / Average exposure time: 1.0 sec. / Average electron dose: 5.0 e/Å2
Details: This is a representative tomogram of many samples and grids imaged.

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Image processing

Final reconstructionAlgorithm: BACK PROJECTION / Software - Name: IMOD (ver. 4.3) / Number images used: 51

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