Journal: Prog Biophys Mol Biol / Year: 2021 Title: Localized reconstruction in Scipion expedites the analysis of symmetry mismatches in cryo-EM data. Authors: Vahid Abrishami / Serban L Ilca / Josue Gomez-Blanco / Ilona Rissanen / José Miguel de la Rosa-Trevín / Vijay S Reddy / José-Maria Carazo / Juha T Huiskonen / Abstract: Technological advances in transmission electron microscopes and detectors have turned cryogenic electron microscopy (cryo-EM) into an essential tool for structural biology. A commonly used cryo-EM ...Technological advances in transmission electron microscopes and detectors have turned cryogenic electron microscopy (cryo-EM) into an essential tool for structural biology. A commonly used cryo-EM data analysis method, single particle analysis, averages hundreds of thousands of low-dose images of individual macromolecular complexes to determine a density map of the complex. The presence of symmetry in the complex is beneficial since each projection image can be assigned to multiple views of the complex. However, data processing that applies symmetry can average out asymmetric features and consequently data analysis methods are required to resolve asymmetric structural features. Scipion is a cryo-EM image processing framework that integrates functions from different image processing packages as plugins. To extend its functionality for handling symmetry mismatches, we present here a Scipion plugin termed LocalRec implementing the localized reconstruction method. When tested on an adenovirus data set, the plugin enables resolving the symmetry-mismatched trimeric fibre bound to the five-fold vertices of the capsid. Furthermore, it improves the structure determination of the icosahedral capsid by dealing with the defocus gradient across the particle. LocalRec is expected to be widely applicable in a range of cryo-EM investigations of flexible and symmetry mismatched complexes.
History
Deposition
May 6, 2020
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Header (metadata) release
Jun 10, 2020
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Map release
Jun 10, 2020
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Update
Mar 24, 2021
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Current status
Mar 24, 2021
Processing site: PDBe / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
EMPIAR-10455 (Title: Single particle cryo-EM dataset of human adenovirus HAdV-D26 Data size: 46.5 Data #1: Particle stacks of human adenovirus HAdV-D26 [picked particles - single frame - processed])
Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 7-38 / Number real images: 2000 / Average exposure time: 7.6 sec. / Average electron dose: 53.0 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron optics
Illumination mode: OTHER / Imaging mode: OTHER / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 22500
Number selected: 521040 Details: A total of 521040 sub-particle images were extracted from 8,684 particle images.
CTF correction
Software - Name: CTFFIND (ver. 4.1.8)
Startup model
Type of model: OTHER / Details: Localized reconstruction of the fibre
Final reconstruction
Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 521040
Initial angle assignment
Type: OTHER / Software - Name: Scipion / Software - details: Localized reconstruction Details: Orientation parameters were calculated by localized reconstruction from the alignment of the capsid
Final angle assignment
Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0) Software - details: custom option was added to relax C5 symmetry
Final 3D classification
Number classes: 3 / Software - Name: RELION (ver. 3.0)
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