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Conformations of Microtubule-Associated Protein Tau Mapped by Fluorescence Resonance Energy Transfer

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Amyloid Proteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 849))

Abstract

The microtubule-associated protein Tau plays a physiological role of stabilizing neuronal microtubules by binding to their lateral surface. Tau belongs to the category of natively unfolded protein as it shows typical features of random coil, as analyzed by various biophysical techniques. In cells, it is subjected to several posttranslational modifications (e.g., phosphorylation, cleavage, ubiquitination, and glycosylation). In neurodegenerative diseases, Tau forms insoluble aggregates called paired helical filaments (PHFs). We have applied fluorescence resonance energy transfer (FRET) to examine the conformations of soluble Tau. We created a series of Tau mutants, each carrying one tryptophan and one cysteine (labeled by IEADANS). This made it possible to measure the distance between these FRET pairs placed in different domains of Tau. This approach enables one to analyze the global folding of soluble Tau and its alteration upon phosphorylation and denaturation.

Sadasivam Jeganathan and Subashchandrabose Chinnathambi contributed equally.

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Abbreviations

CD:

Circular dichroism

DTT:

Dithiothreitol

FDTP-17:

Frontotemporal dementia and parkinsonism linked to chromosome 17

FRET:

Fluorescence resonance energy transfer

FTIR:

Fourier transform infrared

GdnHCl:

Guanidine hydrochloride

IAEDANS:

5-(((2-Idoacetyl) amino) ethyl) amino) naphthalene-1-sulfonic acid

NMR:

Nuclear magnetic resonance spectroscopy

PHFs:

Paired helical filaments

SAXS:

Small-angle X-ray scattering

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Acknowledgments

We thank Bianca Scharnweber and Ilka Lindner for excellent technical assistance, and Dr. Jacek Biernat for advice and discussions on Tau mutations and purification. This project was supported in part by grants from the DFG, MPG (TPC project), V W Foundation (Protein Conformation project), and Breuer Foundation.

Author information

Authors and Affiliations

  1. Max-Planck-Unit for Structural Molecular Biology c/o DESY, Hamburg, Germany

    Sadasivam Jeganathan, Eva-Maria Mandelkow & Eckhard Mandelkow

  2. Department of Experimental Oncology, IFOM-IEO, Campus, Milan, Italy

    Sadasivam Jeganathan

  3. Max Planck Unit for Structural Molecular Biology c/o DESY, Hamburg, Germany

    Subashchandrabose Chinnathambi

  4. DZNE, German Center for Neurodegenerative Diseases c/o CAESAR, Hamburg, Germany

    Eva-Maria Mandelkow & Eckhard Mandelkow

Authors
  1. Sadasivam Jeganathan
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  2. Subashchandrabose Chinnathambi
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  3. Eva-Maria Mandelkow
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  4. Eckhard Mandelkow
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Corresponding author

Correspondence to Eckhard Mandelkow .

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Editors and Affiliations

  1. School of Medicine, Dept. Psychiatry, New York University, First Ave. 550, New York, 10016, New York, USA

    Einar M. Sigurdsson

  2. Centro Nacional de Microbiología, Instituto de Salud Carlos III, Carretera Majadahonda-Pozuelo Km 2, Madrid, 28220, Spain

    Miguel Calero

  3. Inst. Química Física, Rocasolano, CSIC Madrid, C/ Serrano 119, Madrid, 28006, Spain

    María Gasset

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Jeganathan, S., Chinnathambi, S., Mandelkow, EM., Mandelkow, E. (2012). Conformations of Microtubule-Associated Protein Tau Mapped by Fluorescence Resonance Energy Transfer. In: Sigurdsson, E., Calero, M., Gasset, M. (eds) Amyloid Proteins. Methods in Molecular Biology, vol 849. Humana Press. https://doi.org/10.1007/978-1-61779-551-0_7

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  • DOI: https://doi.org/10.1007/978-1-61779-551-0_7

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-550-3

  • Online ISBN: 978-1-61779-551-0

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