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- EMDB-56606: Cryo-tomogram of COPI and COPII vesicles and buds from FIB-milled... -

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Basic information

Entry
Database: EMDB / ID: EMD-56606
TitleCryo-tomogram of COPI and COPII vesicles and buds from FIB-milled RPE-1 cells.
Map dataTomogram showing COPI and COPII vesicles and buds derived from plunge frozen, FIB-milled, Halo_Sec23A RPE-1 cells.
Sample
  • Organelle or cellular component: COPI and COPII coats in Halo_Sec23A RPE-1 cells.
KeywordsIntracellular Coat / Human / COPII / COPI / TRANSPORT PROTEIN
Biological speciesHomo sapiens (human)
Methodelectron tomography / cryo EM / negative staining
AuthorsDownes KW / Zanetti G / Nans A
Funding supportEuropean Union, 1 items
OrganizationGrant numberCountry
European Research Council (ERC)ERC-StG-2019 grant 852915European Union
CitationJournal: biorxiv
Title: Multi-scale Molecular Imaging of Human Cells reveals COPI and COPII Vesicles at ER Exit Sites
Authors: Downes KW / Flood J / Nans A / VanderVerren S / Audhya A / Zanetti G
History
DepositionFeb 6, 2026-
Header (metadata) releaseApr 15, 2026-
Map releaseApr 15, 2026-
UpdateApr 15, 2026-
Current statusApr 15, 2026Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_56606.png

Downloads & links

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Map

FileDownload / File: emd_56606.map.gz / Format: CCP4 / Size: 1.3 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTomogram showing COPI and COPII vesicles and buds derived from plunge frozen, FIB-milled, Halo_Sec23A RPE-1 cells.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
10 Å/pix.
x 448 pix.
= 4480. Å		10 Å/pix.
x 896 pix.
= 8960. Å		10 Å/pix.
x 896 pix.
= 8960. Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 10 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-0.16640995 - 0.24021563
Average (Standard dev.)-0.00004516539 (±0.0049080825)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-431
Dimensions896896448
Spacing896896448
CellA: 8960.0 Å / B: 8960.0 Å / C: 4480.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : COPI and COPII coats in Halo_Sec23A RPE-1 cells.

EntireName: COPI and COPII coats in Halo_Sec23A RPE-1 cells.
Components
  • Organelle or cellular component: COPI and COPII coats in Halo_Sec23A RPE-1 cells.

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Supramolecule #1: COPI and COPII coats in Halo_Sec23A RPE-1 cells.

SupramoleculeName: COPI and COPII coats in Halo_Sec23A RPE-1 cells. / type: organelle_or_cellular_component / ID: 1 / Parent: 0
Details: Halo tag added to endogenous site of Sec23A in RPE-1 cells, used to target lamella production by FIB-milling and tomograms positioning.
Source (natural)Organism: Homo sapiens (human) / Strain: Halo_Sec23A RPE-1 / Organelle: ERES

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processingelectron tomography
Aggregation state3D array

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Sample preparation

BufferpH: 7.4 / Component - Concentration: 1.0 AU / Component - Formula: DMEM/F12
Component - Name: DMEM/F12, 10% FBS, L-glutamine, 1% Pen/Strep
StainingType: NONE / Material: Oregon Green-HaloTag Ligand
Details: Cells stained with 1 uL/mL Oregon Green-HaloTag Ligand
GridModel: Quantifoil R2/2 / Material: MOLYBDENUM / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Details: Tergeo-EM (PIE scientific)
VitrificationCryogen name: ETHANE / Chamber humidity: 70 % / Chamber temperature: 310 K / Instrument: LEICA EM GP
SectioningFocused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 / Focused ion beam - Current: 1 / Focused ion beam - Duration: 3000 / Focused ion beam - Temperature: 81 K / Focused ion beam - Initial thickness: 800 / Focused ion beam - Final thickness: 160
Focused ion beam - Details: Lamella positions, eucentric height and milling angles were automatically calculated in AutoTEM 2.4 (Thermo Fisher). Final lamella placement was guided by the site of ...Focused ion beam - Details: Lamella positions, eucentric height and milling angles were automatically calculated in AutoTEM 2.4 (Thermo Fisher). Final lamella placement was guided by the site of interest/fiducial pair that was added in MAPS and identified in the FIB image. The length and position of the lamella was optimized for each cell. Lamellae were produced using a Gallium beam, operating at 30 kV, in a stepwise manner starting at a beam current of 1.0 nA for rough milling and decreasing to 0.5 nA and 0.3 nA for medium and fine milling. Thinning was then performed in two stages with a beam current of first 50 pA, and then 30 pA. Target milling angles ranged from 9 to 15 and the target lamella thickness was set between 150 and 250 nm.. The value given for _em_focused_ion_beam.instrument is Aquilos2. This is not in a list of allowed values {'DB235', 'OTHER'} so OTHER is written into the XML file.

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K2 BASE (4k x 4k) / Average electron dose: 3.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 8.0 µm / Nominal defocus min: 8.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionSoftware - Name: Warp / Number images used: 42
CTF correctionSoftware - Name: Warp / Details: WARP pipeline / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION

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