ELECTRON CRYSTALLOGRAPHY / electron crystallography / AB INITIO PHASING / cryo EM / Resolution: 1.1 Å
Model details
Protofilament structure of Amyloid-beta 20-34 with the age-associated post-translational ...Protofilament structure of Amyloid-beta 20-34 with the age-associated post-translational modification of aspartate isomerization at position 23
National Institutes of Health/National Center for Research Resources (NIH/NCRR)
5T32GM008496
United States
National Science Foundation (NSF, United States)
MCB-1714569
United States
National Institutes of Health/National Center for Research Resources (NIH/NCRR)
GM-007185
United States
Howard Hughes Medical Institute (HHMI)
United States
Citation
Journal: Nat Commun / Year: 2019 Title: Structure of amyloid-β (20-34) with Alzheimer's-associated isomerization at Asp23 reveals a distinct protofilament interface. Authors: Rebeccah A Warmack / David R Boyer / Chih-Te Zee / Logan S Richards / Michael R Sawaya / Duilio Cascio / Tamir Gonen / David S Eisenberg / Steven G Clarke / Abstract: Amyloid-β (Aβ) harbors numerous posttranslational modifications (PTMs) that may affect Alzheimer's disease (AD) pathogenesis. Here we present the 1.1 Å resolution MicroED structure of an Aβ 20- ...Amyloid-β (Aβ) harbors numerous posttranslational modifications (PTMs) that may affect Alzheimer's disease (AD) pathogenesis. Here we present the 1.1 Å resolution MicroED structure of an Aβ 20-34 fibril with and without the disease-associated PTM, L-isoaspartate, at position 23 (L-isoAsp23). Both wild-type and L-isoAsp23 protofilaments adopt β-helix-like folds with tightly packed cores, resembling the cores of full-length fibrillar Aβ structures, and both self-associate through two distinct interfaces. One of these is a unique Aβ interface strengthened by the isoaspartyl modification. Powder diffraction patterns suggest a similar structure may be adopted by protofilaments of an analogous segment containing the heritable Iowa mutation, Asp23Asn. Consistent with its early onset phenotype in patients, Asp23Asn accelerates aggregation of Aβ 20-34, as does the L-isoAsp23 modification. These structures suggest that the enhanced amyloidogenicity of the modified Aβ segments may also reduce the concentration required to achieve nucleation and therefore help spur the pathogenesis of AD.
Mass: 18.015 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Formula: H2O
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Experimental details
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Experiment
Experiment
Method: ELECTRON CRYSTALLOGRAPHY
EM experiment
Aggregation state: 3D ARRAY / 3D reconstruction method: electron crystallography
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Sample preparation
Component
Name: crystal of amyloid-beta residues 20-34 wild type / Type: COMPLEX / Entity ID: #1 / Source: MULTIPLE SOURCES
Molecular weight
Value: 6.21 kDa/nm / Experimental value: NO
Source (natural)
Organism: Homo sapiens (human)
EM crystal formation
Instrument: Varioscan plate reader / Atmosphere: air / Details: shaken at 1200 rpm / Temperature: 310 K / Time: 30 HOUR
Buffer solution
pH: 7.5
Buffer component
ID
Conc.
Name
Formula
Buffer-ID
1
water
1
2
50mM
trisbase
1
3
150mM
NaCl
1
4
1 %
dimethylsulfoxide
1
Specimen
Conc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: This sample is a crystal.
Specimen support
Details: 30 seconds on each side / Grid type: Quantifoil, UltrAuFoil, R1.2/1.3
Vitrification
Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Chamber temperature: 100 K
Crystal
Density Matthews: 1.5 Å3/Da / Density % sol: 18.21 %
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Data collection
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
Microscopy
Model: FEI TALOS ARCTICA
Electron gun
Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lens
Mode: DIFFRACTION / Alignment procedure: BASIC
Specimen holder
Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 100 K / Temperature (min): 100 K
Image recording
Average exposure time: 3 sec. / Electron dose: 0.01 e/Å2 / Film or detector model: FEI CETA (4k x 4k) / Num. of diffraction images: 404 / Num. of grids imaged: 2
Image scans
Width: 2048 / Height: 2048
EM diffraction
Camera length: 1050 mm
EM diffraction shell
Resolution: 1→1.05 Å / Fourier space coverage: 41.2 % / Multiplicity: 3.09 / Num. of structure factors: 315 / Phase residual: 0.1 °
EM diffraction stats
Fourier space coverage: 85.2 % / High resolution: 1.1 Å / Num. of intensities measured: 23638 / Num. of structure factors: 3546 / Phase error: 22.7 ° / Phase residual: 0.1 ° / Phase error rejection criteria: 0.1 / Rmerge: 0.189 / Rsym: 0.189
Diffraction source
Source: ELECTRON MICROSCOPE / Type: FEI TALOS ARCTICA / Wavelength: 0.0251 Å
Detector
Type: FEI CETA (4k x 4k) / Detector: CMOS DETECTOR / Date: Mar 21, 2019
∠α: 90 ° / ∠β: 111.1 ° / ∠γ: 90 ° / A: 33.17 Å / B: 4.78 Å / C: 30.33 Å / Space group name: P21 / Space group num: 4
CTF correction
Type: NONE
3D reconstruction
Resolution method: DIFFRACTION PATTERN/LAYERLINES / Symmetry type: 3D CRYSTAL
Atomic model building
Protocol: OTHER / Space: RECIPROCAL / Target criteria: maximum liklihood
Refinement
Method to determine structure: AB INITIO PHASING / Resolution: 1.1→7.737 Å / SU ML: 0.13 / Cross valid method: THROUGHOUT / σ(F): 1.41 / Phase error: 22.68
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