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- EMDB-19430: Low-dose cryo-electron ptychographic reconstruction of phi92-shea... -

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Basic information

Entry
Database: EMDB / ID: EMD-19430
TitleLow-dose cryo-electron ptychographic reconstruction of phi92-sheath recorded with CSA of 5.1 mrad
Map datasharpened map
Sample
  • Virus: Staphylococcus phage 92 (virus)
Keywordsphi92 / phage / VIRUS
Biological speciesStaphylococcus phage 92 (virus)
Methodhelical reconstruction / cryo EM / Resolution: 8.4 Å
AuthorsMohammed I / Stalhberg H / Kucukoglu B
Funding support Switzerland, 1 items
OrganizationGrant numberCountry
Swiss National Science Foundation200021_200628 Switzerland
CitationJournal: Nat Commun / Year: 2024
Title: Low-dose cryo-electron ptychography of proteins at sub-nanometer resolution.
Authors: Berk Küçükoğlu / Inayathulla Mohammed / Ricardo C Guerrero-Ferreira / Stephanie M Ribet / Georgios Varnavides / Max Leo Leidl / Kelvin Lau / Sergey Nazarov / Alexander Myasnikov / ...Authors: Berk Küçükoğlu / Inayathulla Mohammed / Ricardo C Guerrero-Ferreira / Stephanie M Ribet / Georgios Varnavides / Max Leo Leidl / Kelvin Lau / Sergey Nazarov / Alexander Myasnikov / Massimo Kube / Julika Radecke / Carsten Sachse / Knut Müller-Caspary / Colin Ophus / Henning Stahlberg /
Abstract: Cryo-transmission electron microscopy (cryo-EM) of frozen hydrated specimens is an efficient method for the structural analysis of purified biological molecules. However, cryo-EM and cryo-electron ...Cryo-transmission electron microscopy (cryo-EM) of frozen hydrated specimens is an efficient method for the structural analysis of purified biological molecules. However, cryo-EM and cryo-electron tomography are limited by the low signal-to-noise ratio (SNR) of recorded images, making detection of smaller particles challenging. For dose-resilient samples often studied in the physical sciences, electron ptychography - a coherent diffractive imaging technique using 4D scanning transmission electron microscopy (4D-STEM) - has recently demonstrated excellent SNR and resolution down to tens of picometers for thin specimens imaged at room temperature. Here we apply 4D-STEM and ptychographic data analysis to frozen hydrated proteins, reaching sub-nanometer resolution 3D reconstructions. We employ low-dose cryo-EM with an aberration-corrected, convergent electron beam to collect 4D-STEM data for our reconstructions. The high frame rate of the electron detector allows us to record large datasets of electron diffraction patterns with substantial overlaps between the interaction volumes of adjacent scan positions, from which the scattering potentials of the samples are iteratively reconstructed. The reconstructed micrographs show strong SNR enabling the reconstruction of the structure of apoferritin protein at up to 5.8 Å resolution. We also show structural analysis of the Phi92 capsid and sheath, tobacco mosaic virus, and bacteriorhodopsin at slightly lower resolutions.
History
DepositionJan 17, 2024-
Header (metadata) releaseFeb 28, 2024-
Map releaseFeb 28, 2024-
UpdateOct 2, 2024-
Current statusOct 2, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19430.png

Downloads & links

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Map

FileDownload / File: emd_19430.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationsharpened map
Projections & slices

Image control

Size
Brightness
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.92 Å/pix.
x 300 pix.
= 574.53 Å		1.92 Å/pix.
x 300 pix.
= 574.53 Å		1.92 Å/pix.
x 300 pix.
= 574.53 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.9151 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.45
Minimum - Maximum-1.0327994 - 1.5581485
Average (Standard dev.)0.004188716 (±0.12476664)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 574.52997 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19430_msk_1.map
Projections & Slices
AxesZYX

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Histogram

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Half map: #2

Fileemd_19430_half_map_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_19430_half_map_2.map
Projections & Slices
AxesZYX

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Sample components

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Entire : Staphylococcus phage 92

EntireName: Staphylococcus phage 92 (virus)
Components
  • Virus: Staphylococcus phage 92 (virus)

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Supramolecule #1: Staphylococcus phage 92

SupramoleculeName: Staphylococcus phage 92 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: #1 / NCBI-ID: 2908151 / Sci species name: Staphylococcus phage 92 / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: Yes / Virus empty: No

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV
Detailsphi92 phage

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: DECTRIS ELA (1k x 0.5k) / Average electron dose: 43.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: DIFFRACTION / Nominal defocus max: 1.7 µm / Nominal defocus min: 1.4000000000000001 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Detailspy4DSTEM CryoSparc
Final reconstructionApplied symmetry - Helical parameters - Δz: 35.3 Å
Applied symmetry - Helical parameters - Δ&Phi: 26.3 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 8.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.1) / Number images used: 1600
Startup modelType of model: NONE / Details: ab-initio cylinder
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.1)
FSC plot (resolution estimation)

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