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- EMDB-21248: Subtomogram average of filamentous segments from in vitro aggrega... -

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Basic information

Entry
Database: EMDB / ID: EMD-21248
TitleSubtomogram average of filamentous segments from in vitro aggregates of mutant huntingtin exon 1 (Q51)
Map dataSubtomogram average of filamentous segments from in vitro aggregates of mutant huntingtin exon 1 (Q51)
Sample
  • Complex: Mutant huntingtin exon-1 Q51 filament
Biological speciesPichia kudriavzevii (yeast)
Methodsubtomogram averaging / cryo EM / Resolution: 35.0 Å
AuthorsGalaz-Montoya JG / Shahmoradian SH / Shen K / Frydman J / Chiu W
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS092525 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P41GM103832 United States
CitationJournal: Commun Biol / Year: 2021
Title: Cryo-electron tomography provides topological insights into mutant huntingtin exon 1 and polyQ aggregates.
Authors: Jesús G Galaz-Montoya / Sarah H Shahmoradian / Koning Shen / Judith Frydman / Wah Chiu /
Abstract: Huntington disease (HD) is a neurodegenerative trinucleotide repeat disorder caused by an expanded poly-glutamine (polyQ) tract in the mutant huntingtin (mHTT) protein. The formation and topology of ...Huntington disease (HD) is a neurodegenerative trinucleotide repeat disorder caused by an expanded poly-glutamine (polyQ) tract in the mutant huntingtin (mHTT) protein. The formation and topology of filamentous mHTT inclusions in the brain (hallmarks of HD implicated in neurotoxicity) remain elusive. Using cryo-electron tomography and subtomogram averaging, here we show that mHTT exon 1 and polyQ-only aggregates in vitro are structurally heterogenous and filamentous, similar to prior observations with other methods. Yet, we find filaments in both types of aggregates under ~2 nm in width, thinner than previously reported, and regions forming large sheets. In addition, our data show a prevalent subpopulation of filaments exhibiting a lumpy slab morphology in both aggregates, supportive of the polyQ core model. This provides a basis for future cryoET studies of various aggregated mHTT and polyQ constructs to improve their structure-based modeling as well as their identification in cells without fusion tags.
History
DepositionJan 29, 2020-
Header (metadata) releaseAug 25, 2021-
Map releaseAug 25, 2021-
UpdateAug 25, 2021-
Current statusAug 25, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 3
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21248.png

Downloads & links

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Map

FileDownload / File: emd_21248.map.gz / Format: CCP4 / Size: 2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram average of filamentous segments from in vitro aggregates of mutant huntingtin exon 1 (Q51)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
8.8 Å/pix.
x 80 pix.
= 704. Å		8.8 Å/pix.
x 80 pix.
= 704. Å		8.8 Å/pix.
x 80 pix.
= 704. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 8.8 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 4.0 / Movie #1: 3
Minimum - Maximum-11.72219 - 23.33857
Average (Standard dev.)0.0809325 (±0.9305763)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-40-40-40
Dimensions808080
Spacing808080
CellA=B=C: 704.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z8.88.88.8
M x/y/z808080
origin x/y/z0.0000.0000.000
length x/y/z704.000704.000704.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-40-40-40
NC/NR/NS808080
D min/max/mean-11.72223.3390.081

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Supplemental data

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Sample components

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Entire : Mutant huntingtin exon-1 Q51 filament

EntireName: Mutant huntingtin exon-1 Q51 filament
Components
  • Complex: Mutant huntingtin exon-1 Q51 filament

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Supramolecule #1: Mutant huntingtin exon-1 Q51 filament

SupramoleculeName: Mutant huntingtin exon-1 Q51 filament / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Pichia kudriavzevii (yeast)
Recombinant expressionOrganism: Pichia kudriavzevii (yeast)
Molecular weightTheoretical: 10 kDa/nm

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation state3D array

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Sample preparation

BufferpH: 7
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: NITROGEN / Chamber temperature: 22 K / Instrument: FEI VITROBOT MARK III

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Electron microscopy

MicroscopeJEOL 2100
Image recordingFilm or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Average electron dose: 1.311 e/Å2
Electron beamAcceleration voltage: 100 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD

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Image processing

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 35.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: EMAN2 / Number subtomograms used: 270
ExtractionNumber tomograms: 20 / Number images used: 450 / Method: manual / Software - Name: EMAN2
Final 3D classificationNumber classes: 2 / Avg.num./class: 270 / Software - Name: EMAN2
Details: Of 450 subtomograms picked, the best 60% was kept in the final average.
Final angle assignmentType: OTHER / Software - Name: EMAN2

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