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- EMDB-71113: ExoSloNano: STA on nucleosomes from cryo-FIB-ET -

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Basic information

Entry
Database: EMDB / ID: EMD-71113
TitleExoSloNano: STA on nucleosomes from cryo-FIB-ET
Map dataSubtomogram analysis of 4955 particles from in situ cryo-ET data
Sample
  • Complex: Nucleosomes from RPE1 mH2A/H2AFY-Halo cells treated with Streptolysin O and 1.4 nm-HaloAlexaNanogold-595.
KeywordsSubtomogram analysis of nucleosomes / NUCLEAR PROTEIN
Biological speciesHomo sapiens (human)
Methodsubtomogram averaging / cryo EM / Resolution: 18.0 Å
AuthorsYoung L / Zhou H / Villa E
Funding support United States, 1 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
Citation
Journal: Nat Methods / Year: 2025
Title: ExoSloNano: multimodal nanogold labels for identification of macromolecules in live cells and cryo-electron tomograms.
Authors: Lindsey N Young / Alice Sherrard / Huabin Zhou / Farhaz Shaikh / Joshua Hutchings / Margot Riggi / Mythreyi Narasimhan / W Alexander Flaherty / Eric J Bennett / Michael K Rosen / Antonio J ...Authors: Lindsey N Young / Alice Sherrard / Huabin Zhou / Farhaz Shaikh / Joshua Hutchings / Margot Riggi / Mythreyi Narasimhan / W Alexander Flaherty / Eric J Bennett / Michael K Rosen / Antonio J Giraldez / Elizabeth Villa /
Abstract: In situ cryo-electron microscopy (cryo-EM) enables the direct interrogation of structure-function relationships by resolving macromolecular structures in their native cellular environment. Recent ...In situ cryo-electron microscopy (cryo-EM) enables the direct interrogation of structure-function relationships by resolving macromolecular structures in their native cellular environment. Recent progress in sample preparation, imaging and data processing has enabled the identification and determination of large biomolecular complexes. However, the majority of proteins are of a size that still eludes identification in cellular cryo-EM data, and most proteins exist in low copy numbers. Therefore, novel tools are needed for cryo-EM to identify macromolecules across multiple size scales (from microns to nanometers). Here we introduce nanogold probes for detecting specific proteins using correlative light and electron microscopy, cryo-electron tomography (cryo-ET) and resin-embedded electron microscopy. These nanogold probes can be introduced into live cells, in a manner that preserves intact molecular networks and cell viability. We use this ExoSloNano system to identify both cytoplasmic and nuclear proteins by room-temperature electron microscopy, and resolve associated structures by cryo-ET. By providing high-efficiency protein labeling in live cells and molecular specificity within cryo-ET tomograms, ExoSloNano expands the proteome available to electron microscopy.
#1: Journal: bioRxiv / Year: 2024
Title: ExoSloNano: Multi-Modal Nanogold Tags for identification of Macromolecules in Live Cells & Cryo-Electron Tomograms
Authors: Young LN / Sherrard A / Zhou H / Shaikh F / Hutchings J / Riggi M / Rosen MK / Giraldez AJ / Villa E
History
DepositionJun 10, 2025-
Header (metadata) releaseNov 19, 2025-
Map releaseNov 19, 2025-
UpdateDec 10, 2025-
Current statusDec 10, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_71113.png

Downloads & links

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Map

FileDownload / File: emd_71113.map.gz / Format: CCP4 / Size: 844.7 KB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram analysis of 4955 particles from in situ cryo-ET data
Projections & slices

Image control

Size
Brightness
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
4 Å/pix.
x 60 pix.
= 240. Å		4 Å/pix.
x 60 pix.
= 240. Å		4 Å/pix.
x 60 pix.
= 240. Å

Surface

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Slices (1/2)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 4 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.08
Minimum - Maximum-0.58691376 - 2.5819871
Average (Standard dev.)0.01932495 (±0.13050084)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions606060
Spacing606060
CellA=B=C: 240.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_71113_msk_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: Unfiltered half-map

Fileemd_71113_half_map_1.map
AnnotationUnfiltered half-map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Unfiltered half-amp

Fileemd_71113_half_map_2.map
AnnotationUnfiltered half-amp
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Nucleosomes from RPE1 mH2A/H2AFY-Halo cells treated with Streptol...

EntireName: Nucleosomes from RPE1 mH2A/H2AFY-Halo cells treated with Streptolysin O and 1.4 nm-HaloAlexaNanogold-595.
Components
  • Complex: Nucleosomes from RPE1 mH2A/H2AFY-Halo cells treated with Streptolysin O and 1.4 nm-HaloAlexaNanogold-595.

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Supramolecule #1: Nucleosomes from RPE1 mH2A/H2AFY-Halo cells treated with Streptol...

SupramoleculeName: Nucleosomes from RPE1 mH2A/H2AFY-Halo cells treated with Streptolysin O and 1.4 nm-HaloAlexaNanogold-595.
type: complex / ID: 1 / Parent: 0 / Details: Human RPE1 mH2A cells
Source (natural)Organism: Homo sapiens (human) / Strain: RPE1 H2AFY-Halo
Molecular weightTheoretical: 200 KDa

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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: 7.2
GridModel: Quantifoil R1/4 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE-PROPANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOCONTINUUM (6k x 4k) / Number real images: 1 / Average electron dose: 4.0 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: 2.0 µm / Nominal magnification: 105000
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: 18.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4) / Number subtomograms used: 4955
ExtractionNumber tomograms: 11 / Number images used: 6603 / Software - Name: RELION (ver. 4)
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Final 3D classificationSoftware - Name: RELION (ver. 4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT

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