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Expression, Biochemistry, and Stabilization with Camel Antibodies of Membrane Proteins: Case Study of the Mouse 5-HT3 Receptor

  • Ghérici Hassaïne
  • Cédric Deluz
  • Luigino Grasso
  • Romain Wyss
  • Ruud Hovius
  • Henning Stahlberg
  • Takashi Tomizaki
  • Aline Desmyter
  • Christophe Moreau
  • Lucie Peclinovska
  • Sonja Minniberger
  • Lamia Mebarki
  • Xiao-Dan Li
  • Horst Vogel
  • Hugues Nury
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1635)

Abstract

There is growing interest in the use of mammalian protein expression systems, and in the use of antibody-derived chaperones, for structural studies. Here, we describe protocols ranging from the production of recombinant membrane proteins in stable inducible cell lines to biophysical characterization of purified membrane proteins in complex with llama antibody domains. These protocols were used to solve the structure of the mouse 5-HT3 serotonin receptor but are of broad applicability for crystallization or cryo-electron microscopy projects.

Key words

Membrane protein Stable cell line VHH Lama antibody Cys-loop receptor 

Notes

Acknowledgments

The work was supported by a grant from the European Research Council (ERC-2014-StG PentaBrain), by the Swiss National Science Foundation, by the Ecole Polytechnique Fédérale de Lausanne, and by the CEA.

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Ghérici Hassaïne
    • 1
  • Cédric Deluz
    • 2
  • Luigino Grasso
    • 2
  • Romain Wyss
    • 2
  • Ruud Hovius
    • 2
  • Henning Stahlberg
    • 3
  • Takashi Tomizaki
    • 4
  • Aline Desmyter
    • 5
  • Christophe Moreau
    • 6
  • Lucie Peclinovska
    • 6
  • Sonja Minniberger
    • 6
  • Lamia Mebarki
    • 1
  • Xiao-Dan Li
    • 7
  • Horst Vogel
    • 2
  • Hugues Nury
    • 6
  1. 1.TheranyxMarseilleFrance
  2. 2.Laboratory of Physical Chemistry of Polymers and MembranesEcole Polytechnique Fédérale de LausanneLausanneSwitzerland
  3. 3.Center for Cellular Imaging and NanoAnalyticsBiozentrum, University of BaselBaselSwitzerland
  4. 4.Swiss Light SourcePaul Scherrer InstituteVilligenSwitzerland
  5. 5.Architecture et Fonction des Macromolécules BiologiquesUnité Mixte de Recherche 7257 Centre National de la Recherche ScientifiqueAix-MarseilleFrance
  6. 6.Univ. Grenoble Alpes, CNRS, CEA, CNRSGrenobleFrance
  7. 7.Laboratory of Biomolecular ResearchPaul Scherrer InstituteVilligenSwitzerland

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