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- EMDB-31004: Heme arrested off-pathway oligomer of alpha-synuclein -

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Basic information

Entry
Database: EMDB / ID: EMD-31004
TitleHeme arrested off-pathway oligomer of alpha-synuclein
Map dataIt is a cryo-EM map of alfa-syneuclein tetramer in presence of heme. alfa-syneuclein monomer was incubated with heme molecule to prepare the sample.
Sample
  • Complex: Heme arrested off-pathway oligomer of alfa-syneuclein
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 10.4 Å
AuthorsDey S / Sil P / Paul S
CitationJournal: Commun Biol / Year: 2021
Title: Conformational distortion in a fibril-forming oligomer arrests alpha-Synuclein fibrillation and minimizes its toxic effects.
Authors: Ritobrita Chakraborty / Sandip Dey / Pallabi Sil / Simanta Sarani Paul / Dipita Bhattacharyya / Anirban Bhunia / Jayati Sengupta / Krishnananda Chattopadhyay /
Abstract: The fibrillation pathway of alpha-Synuclein, the causative protein of Parkinson's disease, encompasses transient, heterogeneous oligomeric forms whose structural understanding and link to toxicity ...The fibrillation pathway of alpha-Synuclein, the causative protein of Parkinson's disease, encompasses transient, heterogeneous oligomeric forms whose structural understanding and link to toxicity are not yet understood. We report that the addition of the physiologically-available small molecule heme at a sub-stoichiometric ratio to either monomeric or aggregated α-Syn, targets a His50 residue critical for fibril-formation and stabilizes the structurally-heterogeneous populations of aggregates into a minimally-toxic oligomeric state. Cryo-EM 3D reconstruction revealed a 'mace'-shaped structure of this monodisperse population of oligomers, which is comparable to a solid-state NMR Greek key-like motif (where the core residues are arranged in parallel in-register sheets with a Greek key topology at the C terminus) that forms the fundamental unit/kernel of protofilaments. Further structural analyses suggest that heme binding induces a distortion in the Greek key-like architecture of the mace oligomers, which impairs their further appending into protofilaments and fibrils. Additionally, our study reports a novel mechanism of prevention as well as reclamation of amyloid fibril formation by blocking an inter-protofilament His50 residue using a small molecule.
History
DepositionFeb 27, 2021-
Header (metadata) releaseApr 14, 2021-
Map releaseApr 14, 2021-
UpdateApr 14, 2021-
Current statusApr 14, 2021Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 10
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 10
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_31004.png

Downloads & links

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Map

FileDownload / File: emd_31004.map.gz / Format: CCP4 / Size: 1.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationIt is a cryo-EM map of alfa-syneuclein tetramer in presence of heme. alfa-syneuclein monomer was incubated with heme molecule to prepare the sample.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.89 Å/pix.
x 72 pix.
= 136.08 Å		1.89 Å/pix.
x 72 pix.
= 136.08 Å		1.89 Å/pix.
x 72 pix.
= 136.08 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.89 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 8.0 / Movie #1: 10
Minimum - Maximum-17.406523 - 43.313667
Average (Standard dev.)0.7616836 (±4.508148)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions727272
Spacing727272
CellA=B=C: 136.08 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.891.891.89
M x/y/z727272
origin x/y/z0.0000.0000.000
length x/y/z136.080136.080136.080
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ256256256
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS727272
D min/max/mean-17.40743.3140.762

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Supplemental data

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Sample components

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Entire : Heme arrested off-pathway oligomer of alfa-syneuclein

EntireName: Heme arrested off-pathway oligomer of alfa-syneuclein
Components
  • Complex: Heme arrested off-pathway oligomer of alfa-syneuclein

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Supramolecule #1: Heme arrested off-pathway oligomer of alfa-syneuclein

SupramoleculeName: Heme arrested off-pathway oligomer of alfa-syneuclein / type: complex / ID: 1 / Parent: 0
Details: To generate off-pathway oligomer, alfa-syneuclein was incubated with heme in shaking (120 rpm)condition for 22 hrs at 37C.
Source (natural)Organism: Homo sapiens (human) / Organ: brain / Tissue: brain tissue / Location in cell: Cytoplasm
Recombinant expressionOrganism: Escherichia coli DH1 (bacteria) / Recombinant strain: bl21 / Recombinant cell: bacterial cell / Recombinant plasmid: pRK172

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.4 / Details: sodium phosphate buffer, pH 7.4
GridModel: Quantifoil / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: LACEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI POLARA 300
Image recordingFilm or detector model: FEI EAGLE (4k x 4k) / Number grids imaged: 2 / Average exposure time: 2.0 sec. / Average electron dose: 20.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: DIFFRACTION
Sample stageSpecimen holder model: GATAN 910 MULTI-SPECIMEN SINGLE TILT CRYO TRANSFER HOLDER
Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: EMAN2 (ver. 2.2)
Startup modelType of model: NONE
Details: Initial model was developed in EMAN2.Then, the initial model was used as reference model for SPIDER 3D Reconstruction.
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 10.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: SPIDER (ver. 23.03) / Number images used: 15770
Initial angle assignmentType: COMMON LINE / Software - Name: EMAN2 (ver. 2.2)
Final angle assignmentType: PROJECTION MATCHING
Projection matching processing - Number reference projections: 83
Software - Name: SPIDER (ver. 23.03)
FSC plot (resolution estimation)

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