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Monitoring Structural Transitions in IDPs by Site-Directed Spin Labeling EPR Spectroscopy

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Intrinsically Disordered Protein Analysis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 895))

Abstract

Electron paramagnetic resonance (EPR) spectroscopy is a technique that specifically detects unpaired electrons. EPR sensitive reporter groups (spin labels or spin probes) can be introduced into biological systems via site-directed spin labeling (SDSL). This is usually accomplished by cysteine-substitution mutagenesis followed by covalent modification of the unique sulfhydryl group with a selective nitroxide reagent. SDSL EPR spectroscopy has been shown to be a sensitive and powerful method to study structural transitions within intrinsically disordered proteins (IDPs). In this chapter, we provide a detailed experimental protocol for this approach and present a few examples of EPR spectral shapes illustrative of various mobility regimes of the spin probe, reflecting different protein topologies.

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Author information

Authors and Affiliations

  1. Architecture et Fonction des Macromolécules Biologiques, UMR 7257 CNRS and Aix-Marseille Université, Marseille, France

    Johnny Habchi, Antoine Gruet & Sonia Longhi

  2. Bioénergétique et Ingénierie des Protéines, UMR 7281 CNRS and Aix-Marseille Université, Marseille, France

    Marlène Martinho, Bruno Guigliarelli & Valérie Belle

Authors
  1. Johnny Habchi
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  2. Marlène Martinho
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  3. Antoine Gruet
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  4. Bruno Guigliarelli
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  5. Sonia Longhi
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  6. Valérie Belle
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Corresponding authors

Correspondence to Sonia Longhi or Valérie Belle .

Editor information

Editors and Affiliations

  1. Department of Molecular Medicine, College of Medicine, University of South Florida, Bruce B. Downs Blvd 12901, Tampa, 33612, Florida, USA

    Vladimir N. Uversky

  2. Dept. Biochemistry & Molecular Biology, Center for Computational Biology &, Indiana University, West 10th Street 410, Indianapolis, 46202, Indiana, USA

    A. Keith Dunker

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Habchi, J., Martinho, M., Gruet, A., Guigliarelli, B., Longhi, S., Belle, V. (2012). Monitoring Structural Transitions in IDPs by Site-Directed Spin Labeling EPR Spectroscopy. In: Uversky, V., Dunker, A. (eds) Intrinsically Disordered Protein Analysis. Methods in Molecular Biology, vol 895. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-927-3_21

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  • DOI: https://doi.org/10.1007/978-1-61779-927-3_21

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-926-6

  • Online ISBN: 978-1-61779-927-3

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