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Using BRET to Detect Ligand-Specific Conformational Changes in Preformed Signalling Complexes

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Book cover Signal Transduction Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 756))

Abstract

Bioluminescence energy transfer (BRET) has become a powerful tool to study protein–protein interactions and conformational changes among interacting proteins. In particular, BRET assays performed in living cells have revealed that heptahelical receptors (7TMRs), heterotrimeric G proteins and their proximal effectors form constitutive signalling complexes. BRET technology has also allowed us to demonstrate that these multimeric protein arrays remain intact throughout initial stages of receptor signalling, thus providing a platform for direct transmission of conformational information from activated receptors to downstream signalling partners. A clear example of the latter are the distinct intermolecular re-arrangements undergone by 7TMRs and G protein subunits following activation of the receptor by different ligands. Here we present protocols describing the type of BRET assay that has been used to reveal the existence of constitutive signalling arrays formed by 7TMRs and proximal signalling partners as well as the ability of complex components to undergo ligand-specific conformational changes.

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Acknowledgements

This work was supported by Discovery Grant from The National Sciences and Engineering Council of Canada (NSERC) to GP.

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Authors and Affiliations

  1. Department of Pharmacology, Faculty of Medicine, University of Montreal, Montreal, QC, Canada

    Nicolas Audet

  2. Centre de Recherche du CHU Ste-Justine, Bureau, Montreal, QC, Canada

    Nicolas Audet & Graciela Piñeyro

  3. Department of Psychiatry, Faculty of Medicine, University of Montreal, Montreal, QC, Canada

    Graciela Piñeyro

Authors
  1. Nicolas Audet
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  2. Graciela Piñeyro
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Editor information

Editors and Affiliations

  1. Div. Endocrinology, Metabolism &, Medical Genetics, Medical University of South Carolina, Jonathan Lucas Street 96, Charleston, 29425, South Carolina, USA

    Louis M. Luttrell

  2. Dept. Physiology & Pharmacology, University of Western Ontario, London, N6A 5C1, Ontario, Canada

    Stephen S. G. Ferguson

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Audet, N., Piñeyro, G. (2011). Using BRET to Detect Ligand-Specific Conformational Changes in Preformed Signalling Complexes. In: Luttrell, L., Ferguson, S. (eds) Signal Transduction Protocols. Methods in Molecular Biology, vol 756. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-160-4_7

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  • DOI: https://doi.org/10.1007/978-1-61779-160-4_7

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-159-8

  • Online ISBN: 978-1-61779-160-4

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