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- EMDB-18874: Cofactor-free Tau 4R2N isoform -

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Basic information

Entry
Database: EMDB / ID: EMD-18874
TitleCofactor-free Tau 4R2N isoform
Map dataAutomatically sharpened and masked map
Sample
  • Complex: In vitro assembled MAPT P-10636 4R2N isoform
    • Protein or peptide: Microtubule-associated protein tau
KeywordsTau / ClearTau / fibrils / aggregation / co-factor-free / heparin-free / PROTEIN FIBRIL
Function / homology
Function and homology information


plus-end-directed organelle transport along microtubule / histone-dependent DNA binding / neurofibrillary tangle assembly / positive regulation of diacylglycerol kinase activity / axonal transport / negative regulation of establishment of protein localization to mitochondrion / neurofibrillary tangle / positive regulation of protein localization to synapse / microtubule lateral binding / tubulin complex ...plus-end-directed organelle transport along microtubule / histone-dependent DNA binding / neurofibrillary tangle assembly / positive regulation of diacylglycerol kinase activity / axonal transport / negative regulation of establishment of protein localization to mitochondrion / neurofibrillary tangle / positive regulation of protein localization to synapse / microtubule lateral binding / tubulin complex / phosphatidylinositol bisphosphate binding / main axon / negative regulation of kinase activity / regulation of long-term synaptic depression / negative regulation of tubulin deacetylation / generation of neurons / rRNA metabolic process / internal protein amino acid acetylation / regulation of chromosome organization / regulation of mitochondrial fission / axonal transport of mitochondrion / intracellular distribution of mitochondria / axon development / central nervous system neuron development / regulation of microtubule polymerization / apolipoprotein binding / microtubule polymerization / lipoprotein particle binding / minor groove of adenine-thymine-rich DNA binding / dynactin binding / negative regulation of mitochondrial membrane potential / glial cell projection / protein polymerization / axolemma / negative regulation of mitochondrial fission / regulation of microtubule polymerization or depolymerization / Caspase-mediated cleavage of cytoskeletal proteins / positive regulation of axon extension / regulation of microtubule cytoskeleton organization / Activation of AMPK downstream of NMDARs / regulation of cellular response to heat / cytoplasmic microtubule organization / positive regulation of protein localization / supramolecular fiber organization / stress granule assembly / regulation of calcium-mediated signaling / axon cytoplasm / somatodendritic compartment / positive regulation of microtubule polymerization / synapse assembly / cellular response to brain-derived neurotrophic factor stimulus / nuclear periphery / phosphatidylinositol binding / cellular response to nerve growth factor stimulus / positive regulation of superoxide anion generation / protein phosphatase 2A binding / regulation of autophagy / astrocyte activation / response to lead ion / microglial cell activation / synapse organization / Hsp90 protein binding / protein homooligomerization / PKR-mediated signaling / regulation of synaptic plasticity / : / memory / SH3 domain binding / microtubule cytoskeleton organization / cytoplasmic ribonucleoprotein granule / cellular response to reactive oxygen species / microtubule cytoskeleton / neuron projection development / cell-cell signaling / protein-folding chaperone binding / single-stranded DNA binding / actin binding / cellular response to heat / protein-macromolecule adaptor activity / double-stranded DNA binding / growth cone / cell body / microtubule binding / sequence-specific DNA binding / microtubule / amyloid fibril formation / dendritic spine / learning or memory / nuclear speck / neuron projection / membrane raft / axon / negative regulation of gene expression / neuronal cell body / DNA damage response / dendrite / protein kinase binding / enzyme binding / mitochondrion / DNA binding
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: 3.1 Å
AuthorsLimorenko G / Tatli M / Kolla R / Nazarov S / Weil MT / Schondorf DC / Geist D / Reinhardt P / Ehrnhoefer DE / Stahlberg H ...Limorenko G / Tatli M / Kolla R / Nazarov S / Weil MT / Schondorf DC / Geist D / Reinhardt P / Ehrnhoefer DE / Stahlberg H / Gasparini L / Lashuel HA
Funding support1 items
OrganizationGrant numberCountry
Other government
CitationJournal: Nat Commun / Year: 2023
Title: Fully co-factor-free ClearTau platform produces seeding-competent Tau fibrils for reconstructing pathological Tau aggregates.
Authors: Galina Limorenko / Meltem Tatli / Rajasekhar Kolla / Sergey Nazarov / Marie-Theres Weil / David C Schöndorf / Daniela Geist / Peter Reinhardt / Dagmar E Ehrnhoefer / Henning Stahlberg / ...Authors: Galina Limorenko / Meltem Tatli / Rajasekhar Kolla / Sergey Nazarov / Marie-Theres Weil / David C Schöndorf / Daniela Geist / Peter Reinhardt / Dagmar E Ehrnhoefer / Henning Stahlberg / Laura Gasparini / Hilal A Lashuel /
Abstract: Tau protein fibrillization is implicated in the pathogenesis of several neurodegenerative diseases collectively known as Tauopathies. For decades, investigating Tau fibrillization in vitro has ...Tau protein fibrillization is implicated in the pathogenesis of several neurodegenerative diseases collectively known as Tauopathies. For decades, investigating Tau fibrillization in vitro has required the addition of polyanions or other co-factors to induce its misfolding and aggregation, with heparin being the most commonly used. However, heparin-induced Tau fibrils exhibit high morphological heterogeneity and a striking structural divergence from Tau fibrils isolated from Tauopathies patients' brains at ultra- and macro-structural levels. To address these limitations, we developed a quick, cheap, and effective method for producing completely co-factor-free fibrils from all full-length Tau isoforms and mixtures thereof. We show that Tau fibrils generated using this ClearTau method - ClearTau fibrils - exhibit amyloid-like features, possess seeding activity in biosensor cells and hiPSC-derived neurons, retain RNA-binding capacity, and have morphological properties and structures more reminiscent of the properties of the brain-derived Tau fibrils. We present the proof-of-concept implementation of the ClearTau platform for screening Tau aggregation-modifying compounds. We demonstrate that these advances open opportunities to investigate the pathophysiology of disease-relevant Tau aggregates and will facilitate the development of Tau pathology-targeting and modifying therapies and PET tracers that can distinguish between different Tauopathies.
History
DepositionNov 10, 2023-
Header (metadata) releaseDec 6, 2023-
Map releaseDec 6, 2023-
SupersessionJan 24, 2024ID: EMD-18874
UpdateJan 24, 2024-
Current statusJan 24, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18874.png

Downloads & links

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Map

FileDownload / File: emd_18874.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationAutomatically sharpened and masked map
Projections & slices

Image control

Size
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.73 Å/pix.
x 384 pix.
= 278.784 Å		0.73 Å/pix.
x 384 pix.
= 278.784 Å		0.73 Å/pix.
x 384 pix.
= 278.784 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.726 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0072
Minimum - Maximum-0.015234422 - 0.031466186
Average (Standard dev.)0.000023706252 (±0.0005815435)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 278.784 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_18874_msk_1.map
Projections & Slices
AxesZYX

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Additional map: Refined map

Fileemd_18874_additional_1.map
AnnotationRefined map
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Half map: Half map1

Fileemd_18874_half_map_1.map
AnnotationHalf map1
Projections & Slices
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Histogram (log scale)

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Half map: Half map2

Fileemd_18874_half_map_2.map
AnnotationHalf map2
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Sample components

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Entire : In vitro assembled MAPT P-10636 4R2N isoform

EntireName: In vitro assembled MAPT P-10636 4R2N isoform
Components
  • Complex: In vitro assembled MAPT P-10636 4R2N isoform
    • Protein or peptide: Microtubule-associated protein tau

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Supramolecule #1: In vitro assembled MAPT P-10636 4R2N isoform

SupramoleculeName: In vitro assembled MAPT P-10636 4R2N isoform / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: ClearTau 4R2N isoform fibrils, PK-digested post-fibrillization
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 9 kDa/nm

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Macromolecule #1: Microtubule-associated protein tau

MacromoleculeName: Microtubule-associated protein tau / type: protein_or_peptide / ID: 1
Details: ClearTau full-length Tau 4R2N isoform fibrils, PK-digested post-fibrillization
Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 45.919871 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MAEPRQEFEV MEDHAGTYGL GDRKDQGGYT MHQDQEGDTD AGLKESPLQT PTEDGSEEPG SETSDAKSTP TAEDVTAPLV DEGAPGKQA AAQPHTEIPE GTTAEEAGIG DTPSLEDEAA GHVTQARMVS KSKDGTGSDD KKAKGADGKT KIATPRGAAP P GQKGQANA ...String:
MAEPRQEFEV MEDHAGTYGL GDRKDQGGYT MHQDQEGDTD AGLKESPLQT PTEDGSEEPG SETSDAKSTP TAEDVTAPLV DEGAPGKQA AAQPHTEIPE GTTAEEAGIG DTPSLEDEAA GHVTQARMVS KSKDGTGSDD KKAKGADGKT KIATPRGAAP P GQKGQANA TRIPAKTPPA PKTPPSSGEP PKSGDRSGYS SPGSPGTPGS RSRTPSLPTP PTREPKKVAV VRTPPKSPSS AK SRLQTAP VPMPDLKNVK SKIGSTENLK HQPGGGKVQI INKKLDLSNV QSKCGSKDNI KHVPGGGSVQ IVYKPVDLSK VTS KCGSLG NIHHKPGGGQ VEVKSEKLDF KDRVQSKIGS LDNITHVPGG GNKKIETHKL TFRENAKAKT DHGAEIVYKS PVVS GDTSP RHLSNVSSTG SIDMVDSPQL ATLADEVSAS LAKQGL

UniProtKB: Microtubule-associated protein tau

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.4
Component:
ConcentrationFormulaName
137.0 mMNaClsodium chloride
2.7 mMKClpotasium chloride
4.3 mMNa2HPOdisodium phosphate
GridModel: Quantifoil R2/1 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 23 sec.
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.6 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

Final reconstructionNumber classes used: 1
Applied symmetry - Helical parameters - Δz: 4.72 Å
Applied symmetry - Helical parameters - Δ&Phi: -0.88 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4.0-beta-2) / Number images used: 15869
Startup modelType of model: OTHER / Details: IniModel2D was used for the initial model creation
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 4.0-beta-2)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 33
Output model

PDB-8r3t:
Cofactor-free Tau 4R2N isoform

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