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- EMDB-4566: Cryo-EM reconstruction of heparin-induced 2N3R tau filaments -

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Basic information

Entry
Database: EMDB / ID: EMD-4566
TitleCryo-EM reconstruction of heparin-induced 2N3R tau filaments
Map dataThe helical reconstruction of heparin-induced 2N3R tau filaments
Sample
  • Complex: heparin-induced 2N3R tau filaments
    • Protein or peptide: Isoform Tau-C of Microtubule-associated protein tau
Function / homology
Function and homology information


plus-end-directed organelle transport along microtubule / histone-dependent DNA binding / neurofibrillary tangle / microtubule lateral binding / axonal transport / tubulin complex / positive regulation of protein localization to synapse / phosphatidylinositol bisphosphate binding / generation of neurons / axonal transport of mitochondrion ...plus-end-directed organelle transport along microtubule / histone-dependent DNA binding / neurofibrillary tangle / microtubule lateral binding / axonal transport / tubulin complex / positive regulation of protein localization to synapse / phosphatidylinositol bisphosphate binding / generation of neurons / axonal transport of mitochondrion / axon development / rRNA metabolic process / central nervous system neuron development / regulation of microtubule-based movement / negative regulation of protein localization to mitochondrion / regulation of mitochondrial fission / regulation of chromosome organization / intracellular distribution of mitochondria / minor groove of adenine-thymine-rich DNA binding / lipoprotein particle binding / microtubule polymerization / negative regulation of mitochondrial membrane potential / regulation of microtubule polymerization / dynactin binding / apolipoprotein binding / regulation of calcium-mediated signaling / main axon / glial cell projection / Caspase-mediated cleavage of cytoskeletal proteins / regulation of microtubule polymerization or depolymerization / neurofibrillary tangle assembly / negative regulation of mitochondrial fission / protein polymerization / positive regulation of axon extension / axolemma / positive regulation of microtubule polymerization / regulation of cellular response to heat / positive regulation of superoxide anion generation / Activation of AMPK downstream of NMDARs / cytoplasmic microtubule organization / cellular response to brain-derived neurotrophic factor stimulus / regulation of long-term synaptic depression / axon cytoplasm / positive regulation of protein localization / supramolecular fiber organization / synapse assembly / somatodendritic compartment / astrocyte activation / nuclear periphery / phosphatidylinositol binding / protein phosphatase 2A binding / stress granule assembly / enzyme inhibitor activity / SH3 domain binding / cellular response to reactive oxygen species / regulation of microtubule cytoskeleton organization / memory / microglial cell activation / cellular response to nerve growth factor stimulus / regulation of synaptic plasticity / Hsp90 protein binding / regulation of autophagy / microtubule cytoskeleton organization / synapse organization / protein homooligomerization / PKR-mediated signaling / response to lead ion / neuron projection development / cytoplasmic ribonucleoprotein granule / microtubule cytoskeleton / cell-cell signaling / cellular response to heat / single-stranded DNA binding / actin binding / protein-folding chaperone binding / growth cone / double-stranded DNA binding / cell body / microtubule / sequence-specific DNA binding / amyloid fibril formation / microtubule binding / dendritic spine / learning or memory / protein-macromolecule adaptor activity / neuron projection / membrane raft / negative regulation of gene expression / axon / neuronal cell body / DNA damage response / dendrite / protein kinase binding / enzyme binding / mitochondrion / DNA binding / RNA binding / extracellular region / identical protein binding / nucleus
Similarity search - Function
Microtubule-associated protein Tau / Microtubule associated protein, tubulin-binding repeat / Tau and MAP protein, tubulin-binding repeat / Tau and MAP proteins tubulin-binding repeat signature. / Tau and MAP proteins tubulin-binding repeat profile. / :
Similarity search - Domain/homology
Microtubule-associated protein tau
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodhelical reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsZhang W / Falcon B / Murzin AG / Fan J / Crowther RA / Goedert M / Scheres SHW
Funding support United Kingdom, 2 items
OrganizationGrant numberCountry
Medical Research Council (United Kingdom)MC_UP_A025_1013 United Kingdom
Medical Research Council (United Kingdom)MC_U105184291 United Kingdom
CitationJournal: Elife / Year: 2019
Title: Heparin-induced tau filaments are polymorphic and differ from those in Alzheimer's and Pick's diseases.
Authors: Wenjuan Zhang / Benjamin Falcon / Alexey G Murzin / Juan Fan / R Anthony Crowther / Michel Goedert / Sjors Hw Scheres /
Abstract: Assembly of microtubule-associated protein tau into filamentous inclusions underlies a range of neurodegenerative diseases. Tau filaments adopt different conformations in Alzheimer's and Pick's ...Assembly of microtubule-associated protein tau into filamentous inclusions underlies a range of neurodegenerative diseases. Tau filaments adopt different conformations in Alzheimer's and Pick's diseases. Here, we used cryo- and immuno- electron microscopy to characterise filaments that were assembled from recombinant full-length human tau with four (2N4R) or three (2N3R) microtubule-binding repeats in the presence of heparin. 2N4R tau assembles into multiple types of filaments, and the structures of three types reveal similar 'kinked hairpin' folds, in which the second and third repeats pack against each other. 2N3R tau filaments are structurally homogeneous, and adopt a dimeric core, where the third repeats of two tau molecules pack in a parallel manner. The heparin-induced tau filaments differ from those of Alzheimer's or Pick's disease, which have larger cores with different repeat compositions. Our results illustrate the structural versatility of amyloid filaments, and raise questions about the relevance of in vitro assembly.
History
DepositionJan 24, 2019-
Header (metadata) releaseFeb 20, 2019-
Map releaseFeb 20, 2019-
UpdateNov 25, 2020-
Current statusNov 25, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.017
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.017
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6qjq
  • Surface level: 0.017
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6qjq
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4566.png

Downloads & links

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Map

FileDownload / File: emd_4566.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThe helical reconstruction of heparin-induced 2N3R tau filaments
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.04 Å/pix.
x 256 pix.
= 266.24 Å		1.04 Å/pix.
x 256 pix.
= 266.24 Å		1.04 Å/pix.
x 256 pix.
= 266.24 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.04 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.017 / Movie #1: 0.017
Minimum - Maximum-0.018145105 - 0.060762867
Average (Standard dev.)0.00106258 (±0.0033862293)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 266.24 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.041.041.04
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z266.240266.240266.240
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ320320320
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0180.0610.001

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Supplemental data

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Mask #1

Fileemd_4566_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: The other half map of helical reconstruction of...

Fileemd_4566_half_map_1.map
AnnotationThe other half map of helical reconstruction of heparin-induced 2N3R tau filaments
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map of helical reconstruction of heparin-induced 2N3R...

Fileemd_4566_half_map_2.map
AnnotationHalf map of helical reconstruction of heparin-induced 2N3R tau filaments
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : heparin-induced 2N3R tau filaments

EntireName: heparin-induced 2N3R tau filaments
Components
  • Complex: heparin-induced 2N3R tau filaments
    • Protein or peptide: Isoform Tau-C of Microtubule-associated protein tau

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Supramolecule #1: heparin-induced 2N3R tau filaments

SupramoleculeName: heparin-induced 2N3R tau filaments / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Recombinant 2N3R tau protein was induced into filaments by adding heparin
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant strain: BL21(DE3) / Recombinant plasmid: pRK172
Molecular weightTheoretical: 42.6 kDa/nm

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Macromolecule #1: Isoform Tau-C of Microtubule-associated protein tau

MacromoleculeName: Isoform Tau-C of Microtubule-associated protein tau / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MAEPRQEFEV MEDHAGTYGL GDRKDQGGYT MHQDQEGDTD AGLKESPLQT PTEDGSEEPG SETSDAKST PTAEDVTAPL VDEGAPGKQA AAQPHTEIPE GTTAEEAGIG DTPSLEDEAA G HVTQARMV SKSKDGTGSD DKKAKGADGK TKIATPRGAA PPGQKGQANA ...String:
MAEPRQEFEV MEDHAGTYGL GDRKDQGGYT MHQDQEGDTD AGLKESPLQT PTEDGSEEPG SETSDAKST PTAEDVTAPL VDEGAPGKQA AAQPHTEIPE GTTAEEAGIG DTPSLEDEAA G HVTQARMV SKSKDGTGSD DKKAKGADGK TKIATPRGAA PPGQKGQANA TRIPAKTPPA PK TPPSSGE PPKSGDRSGY SSPGSPGTPG SRSRTPSLPT PPTREPKKVA VVRTPPKSPS SAK SRLQTA PVPMPDLKNV KSKIGSTENL KHQPGGGKVQ IVYKPVDLSK VTSKCGSLGN IHHK PGGGQ VEVKSEKLDF KDRVQSKIGS LDNITHVPGG GNKKIETHKL TFRENAKAKT DHGAE IVYK SPVVSGDTSP RHLSNVSSTG SIDMVDSPQL ATLADEVSAS LAKQGL

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

Concentration2.0 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
0.02 mol/LTristris(hydroxymethyl)aminomethane
0.1 mol/LNaClsodium chloride

Details: 20 mM Tris, pH 7.4, 100mM NaCl
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 10.0 nm / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: Blot force: -12 ; Blot time: 4s.
DetailsRecombinant tau protein was induced into filaments by incubation with heparin at 37 degree celsius for 3 days

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Digitization - Sampling interval: 5.0 µm / Digitization - Frames/image: 1-44 / Number grids imaged: 1 / Number real images: 2051 / Average exposure time: 11.0 sec. / Average electron dose: 1.134 e/Å2
Details: Images were collected in movie-mode at 44 frames every 11 seconds
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsMovie frames were gain-corrected, aligned, dose weighted and then summed into a single micrograph using MOTIONCOR2 (Zheng et al., 2017)
Final reconstructionNumber classes used: 1
Applied symmetry - Helical parameters - Δz: 4.7 Å
Applied symmetry - Helical parameters - Δ&Phi: -1.05 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0)
Details: For masked FSC calculation, phase randomisation was done at 6.1 A, where the FSC of the unmasked map was 0.85.
Number images used: 149909
CTF correctionSoftware - Name: CTFFIND (ver. 4.1)
Details: Aligned, non-dose-weighted micrographs were used to estimate the contrast transfer function (CTF) using CTFFIND4.1
Segment selectionNumber selected: 788359 / Software - Name: RELION (ver. 3.0) / Details: Manually picked
Startup modelType of model: OTHER
Details: An initial 3D reference was reconstructed from the 2D class averages de novo.
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 3.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

DetailsA stack of three consecutive monomers was refined to preserve nearest-neighbour interactions for the middle chain. Side-chain clashes were detected using MOLPROBITY, and corrected by iterative cycles of real-space refinement in COOT and Fourier-space refinement in REFMAC and PHENIX. For each refined structure, separate model refinements were performed against a single half-map, and the resulting model was compared to the other half-map to confirm the absence of overfitting.
RefinementSpace: RECIPROCAL / Protocol: AB INITIO MODEL / Overall B value: 95.9 / Target criteria: Fourier shell correlation
Output model

PDB-6qjq:
Cryo-EM structure of heparin-induced 2N3R tau filaments

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