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    Yorodumi
    - EMDB-2099: Structure of ER membrane associated ribosomes in situ -

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    Basic information

    Entry
    Database: EMDB / ID: 2099
    TitleStructure of ER membrane associated ribosomes in situ
    Keywords80S ribosome / translocon / mammalian / ER membrane
    SampleER membrane associated ribosome
    SourceCanis lupus familiaris / mammal / Dog /
    Map dataReconstruction of ER membrane associated ribosome from canine pancreatic microsomes
    Methodsubtomogram averaging, at 31 A resolution
    AuthorsPfeffer S / Brandt F / Hrabe T / Lang S / Eibauer M / Zimmermann R / Foerster F
    CitationStructure, 2012, 20, 1508-1518

    Structure, 2012, 20, 1508-1518 StrPapers
    Structure and 3D arrangement of endoplasmic reticulum membrane-associated ribosomes.
    Stefan Pfeffer / Florian Brandt / Thomas Hrabe / Sven Lang / Matthias Eibauer / Richard Zimmermann / Friedrich Förster

    DateDeposition: May 14, 2012 / Header (metadata) release: Jul 25, 2012 / Map release: Jul 25, 2012 / Last update: May 14, 2012

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    Structure visualization

    Movie
    • Surface view with section colored by density value
    • Surface level: 0.00895
    • Imaged by UCSF CHIMERA
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    • Surface view colored by height
    • Surface level: 0.00895
    • Imaged by UCSF CHIMERA
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    Map

    Fileemd_2099.map.gz (map file in CCP4 format, 8193 KB)
    Projections & slicesSize of images:
    AxesZ (Sec.)Y (Row.)X (Col.)
    128 pix
    4.68 A/pix
    = 599.04 A
    128 pix
    4.68 A/pix
    = 599.04 A
    128 pix
    4.68 A/pix
    = 599.04 A

    Surface

    Projections

    Slices (1/3)

    Slices (1/2)

    Slices (2/3)

    Images are generated by Spider package.

    Voxel sizeX=Y=Z: 4.68 A
    Density
    Contour Level:0.00895 (by author), 0.00895 (movie #1):
    Minimum - Maximum-0.05852651 - 0.08826262
    Average (Standard dev.)0.00050413 (0.007813)
    Details

    EMDB XML:

    Space Group Number1
    Map Geometry
    Axis orderXYZ
    Dimensions128128128
    Origin000
    Limit127127127
    Spacing128128128
    CellA=B=C: 599.04 A
    Alpha=beta=gamma: 90 deg.

    CCP4 map header:

    modeImage stored as Reals
    A/pix X/Y/Z4.684.684.68
    M x/y/z128128128
    origin x/y/z0.0000.0000.000
    length x/y/z599.040599.040599.040
    alpha/beta/gamma90.00090.00090.000
    start NX/NY/NZ000
    NX/NY/NZ128128168
    MAP C/R/S123
    start NC/NR/NS000
    NC/NR/NS128128128
    D min/max/mean-0.0590.0880.001

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    Supplemental data

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    Sample components

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    Entire ER membrane associated ribosome

    EntireName: ER membrane associated ribosome / Details: The sample is embedded into its native membrane / Number of components: 1 / Oligomeric State: ER membrane associated ribosome
    MassTheoretical: 4.5 MDa

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    Component #1: ribosome-eukaryote, Membrane-bound 80S ribosome

    Ribosome-eukaryoteName: Membrane-bound 80S ribosome / Eukaryote: ALL / Recombinant expression: No
    MassTheoretical: 4.5 MDa
    SourceSpecies: Canis lupus familiaris / mammal / Dog /
    Source (natural)Organelle: Endoplasmic reticulum / Location in cell: Endoplasmic reticulum / Organ or tissue: Pancreas

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    Experimental details

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    Sample preparation

    Specimen stateparticle
    Sample solutionSpecimen conc.: 1 mg/ml
    Buffer solution: 5 mM MgCl2, 140 mM KCl, 10 mM Hepes pH 7.4, 1 mM DTT, protease inhibitor
    pH: 7.4
    Support filmlacy carbon
    VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Method: Blot for 3 seconds before plunging

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    Electron microscopy imaging

    ImagingMicroscope: FEI POLARA 300 / Date: Jan 1, 2011
    Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 50 e/A2 / Illumination mode: FLOOD BEAM
    LensCs: 2 mm / Imaging mode: BRIGHT FIELD / Defocus: 4000 nm / Energy filter: Gatan GIF 2002 / Energy window: 0-10 eV
    Specimen HolderModel: OTHER / Tilt Angle: -60 - 60 deg. / Temperature: 85 K ( 80 - 90 K)

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    Image acquisition

    Image acquisitionNumber of digital images: 328 / Bit depth: 12

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    Image processing

    ProcessingMethod: subtomogram averaging
    Details: The particles were selected using PyTom and classified by constrained principal component analysis. Average number of projections used in the 3D reconstructions: 1000. Average number of class averages: 1.
    3D reconstructionAlgorithm: Subtomogram averaging / Software: PyTom / CTF correction: each particle / Resolution: 31 A / Resolution method: FSC 0.5

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