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- EMDB-63526: Subtomogram average of GEM-mCherry-nanobody labeled EGFR on A549 ... -

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Basic information

Entry
Database: EMDB / ID: EMD-63526
TitleSubtomogram average of GEM-mCherry-nanobody labeled EGFR on A549 cell membranes
Map dataThe map was processed using ChimeraX's "vop flip" command to ensure it matched the correct handedness.
Sample
  • Organelle or cellular component: Native membranes derived from A549 cells expressing GEM-mCherry-nanobody
KeywordsTag / Agonist / Icosahedron / MEMBRANE PROTEIN
Biological speciesHomo sapiens (human)
Methodsubtomogram averaging / cryo EM / Resolution: 38.0 Å
AuthorsZou T / Zhang J / Zhang Y / Zhang M / Wang H / Pan Y
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)22327808 China
CitationJournal: ACS Chem Biol / Year: 2026
Title: Visualization of EGFR Assembly and Activation Induced by a Protein Nanocage Using Cryo-Electron Tomography.
Authors: Tianyi Zou / Jinrui Zhang / Yaxuan Zhang / Minghan Zhang / Huili Wang / Yangang Pan / Hongda Wang /
Abstract: The epidermal growth factor receptor (EGFR) mediates signal transduction by triggering downstream phosphorylation to regulate cell proliferation. However, the complexity of the cellular environment ...The epidermal growth factor receptor (EGFR) mediates signal transduction by triggering downstream phosphorylation to regulate cell proliferation. However, the complexity of the cellular environment has limited in situ structural investigations of membrane proteins within their native context. Here, we present a proof-of-concept study integrating protein cage labeling with cryo-electron tomography (cryo-ET) to directly visualize receptor assemblies on the native membrane. Using EGFR as a model system, we demonstrate that the protein cage can associate with multiple EGFR molecules, thereby inducing their oligomerization. The distance between neighboring EGFRs within these assemblies was measured to be 7.1 ± 1.2 nm. Furthermore, we validated the functional relevance of this system by showing that protein cage-induced EGFR assemblies were accompanied by enhanced ligand-independent phosphorylation. In summary, our results establish the feasibility of using protein cage-labeling for the induction and in situ structural analysis of membrane protein oligomerization.
History
DepositionFeb 21, 2025-
Header (metadata) releaseFeb 4, 2026-
Map releaseFeb 4, 2026-
UpdateMar 4, 2026-
Current statusMar 4, 2026Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_63526.png

Downloads & links

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Map

FileDownload / File: emd_63526.map.gz / Format: CCP4 / Size: 1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThe map was processed using ChimeraX's "vop flip" command to ensure it matched the correct handedness.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
8.88 Å/pix.
x 64 pix.
= 568.32 Å		8.88 Å/pix.
x 64 pix.
= 568.32 Å		8.88 Å/pix.
x 64 pix.
= 568.32 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 8.88 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.4
Minimum - Maximum-0.8253965 - 2.197665
Average (Standard dev.)0.047900446 (±0.31108937)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions646464
Spacing646464
CellA=B=C: 568.32 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: The map was processed using ChimeraX's "vop flip"...

Fileemd_63526_half_map_1.map
AnnotationThe map was processed using ChimeraX's "vop flip" command to ensure it matched the correct handedness.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: The map was processed using ChimeraX's "vop flip"...

Fileemd_63526_half_map_2.map
AnnotationThe map was processed using ChimeraX's "vop flip" command to ensure it matched the correct handedness.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Native membranes derived from A549 cells expressing GEM-mCherry-n...

EntireName: Native membranes derived from A549 cells expressing GEM-mCherry-nanobody
Components
  • Organelle or cellular component: Native membranes derived from A549 cells expressing GEM-mCherry-nanobody

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Supramolecule #1: Native membranes derived from A549 cells expressing GEM-mCherry-n...

SupramoleculeName: Native membranes derived from A549 cells expressing GEM-mCherry-nanobody
type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 7.4 / Details: phosphate-buffered saline (PBS)
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Digitization - Frames/image: 1-10 / Average electron dose: 3.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 3.4 µm / Nominal magnification: 64000
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: 38.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4.0.1) / Number subtomograms used: 336
ExtractionNumber tomograms: 11 / Number images used: 336 / Reference model: ab initio / Method: manually picked particles / Software - Name: Warp (ver. 1.1.0)
Details: IsoNet-corrected tomograms were used for particle picking
CTF correctionSoftware - Name: Warp (ver. 1.1.0)
Details: Per-particle ctf estimation was performed with Warp
Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Final angle assignmentType: OTHER
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: experimental model
Details: The initial model was generated with direct reconstruction by relion_reconstruction using predetermined orientations
RefinementProtocol: AB INITIO MODEL

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