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- EMDB-50630: mus musculus ribosome from SOLIST lamellas -

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Basic information

Entry
Database: EMDB / ID: EMD-50630
Titlemus musculus ribosome from SOLIST lamellas
Map dataafter M refinement, filtered, sharpened
Sample
  • Cell: mus musculus ribosome from SOLIST lamellas
Keywordsin situ / ribosome / mouse / mus musculus
Biological speciesMus musculus (house mouse)
Methodsubtomogram averaging / Resolution: 8.3 Å
AuthorsErdmann PS / Nguyen HTD / Perone G / Klena N / Vazzana R / Kaluthantrige Don F / Silva M / Sorrentino S / Swuec P / Leroux F ...Erdmann PS / Nguyen HTD / Perone G / Klena N / Vazzana R / Kaluthantrige Don F / Silva M / Sorrentino S / Swuec P / Leroux F / Kalebic N / Coscia F
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nat Methods / Year: 2024
Title: Serialized on-grid lift-in sectioning for tomography (SOLIST) enables a biopsy at the nanoscale.
Authors: Ho Thuy Dung Nguyen / Gaia Perone / Nikolai Klena / Roberta Vazzana / Flaminia Kaluthantrige Don / Malan Silva / Simona Sorrentino / Paolo Swuec / Frederic Leroux / Nereo Kalebic / Francesca ...Authors: Ho Thuy Dung Nguyen / Gaia Perone / Nikolai Klena / Roberta Vazzana / Flaminia Kaluthantrige Don / Malan Silva / Simona Sorrentino / Paolo Swuec / Frederic Leroux / Nereo Kalebic / Francesca Coscia / Philipp S Erdmann /
Abstract: Cryo-focused ion beam milling has substantially advanced our understanding of molecular processes by opening windows into cells. However, applying this technique to complex samples, such as tissues, ...Cryo-focused ion beam milling has substantially advanced our understanding of molecular processes by opening windows into cells. However, applying this technique to complex samples, such as tissues, has presented considerable technical challenges. Here we introduce an innovative adaptation of the cryo-lift-out technique, serialized on-grid lift-in sectioning for tomography (SOLIST), addressing these limitations. SOLIST enhances throughput, minimizes ice contamination and improves sample stability for cryo-electron tomography. It thereby facilitates the high-resolution imaging of a wide range of specimens. We illustrate these advantages on reconstituted liquid-liquid phase-separated droplets, brain organoids and native tissues from the mouse brain, liver and heart. With SOLIST, cellular processes can now be investigated at molecular resolution directly in native tissue. Furthermore, our method has a throughput high enough to render cryo-lift-out a competitive tool for structural biology. This opens new avenues for unprecedented insights into cellular function and structure in health and disease, a 'biopsy at the nanoscale'.
History
DepositionJun 12, 2024-
Header (metadata) releaseSep 11, 2024-
Map releaseSep 11, 2024-
UpdateSep 25, 2024-
Current statusSep 25, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_50630.png

Downloads & links

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Map

FileDownload / File: emd_50630.map.gz / Format: CCP4 / Size: 209.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationafter M refinement, filtered, sharpened
Projections & slices

Image control

Size
Brightness
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
2 Å/pix.
x 380 pix.
= 760. Å		2 Å/pix.
x 380 pix.
= 760. Å		2 Å/pix.
x 380 pix.
= 760. Å

Surface

Projections

Slices (1/3)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 2 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0034
Minimum - Maximum-0.0033790285 - 0.01127715
Average (Standard dev.)0.00003242098 (±0.0006183298)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions380380380
Spacing380380380
CellA=B=C: 760.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_50630_msk_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

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Half map: #1

Fileemd_50630_half_map_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_50630_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram (log scale)

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Sample components

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Entire : mus musculus ribosome from SOLIST lamellas

EntireName: mus musculus ribosome from SOLIST lamellas
Components
  • Cell: mus musculus ribosome from SOLIST lamellas

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Supramolecule #1: mus musculus ribosome from SOLIST lamellas

SupramoleculeName: mus musculus ribosome from SOLIST lamellas / type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Mus musculus (house mouse) / Strain: C57BL/6 N

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Experimental details

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Structure determination

Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 7.5
GridModel: UltrAuFoil R2/2 / Material: GOLD / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE
Details: glow-discharged at 30 mA for 30 s (GloQube Plus Glow discharge system, Quorum Technologies)
DetailsSOLIST lamella preparation of mouse liver from HPF.

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris X / Energy filter - Slit width: 10 eV
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Average electron dose: 3.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.5 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 8.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: Warp (ver. 1.1.0-beta) / Software - details: refinement in Warp/M / Number subtomograms used: 13861
ExtractionNumber tomograms: 43 / Number images used: 43000 / Method: template matching / Software - Name: Warp (ver. 1.1.0-beta) / Details: template matching with PyTOM (GPU)
Final 3D classificationSoftware - Name: RELION (ver. 4.0.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: Warp (ver. 1.1.0-beta)
FSC plot (resolution estimation)

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