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Biochemical Characterization of GPCR–G Protein Complex Formation

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Structure and Function of Membrane Proteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2302))

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Abstract

The complex of G protein-coupled receptors (GPCR) and G proteins is the core assembly in GPCR signaling in eukaryotes. With the recent development of cryo-electron microscopy, there has been a rapid growth in structures of GPCR–G protein complexes solved to near-atomic resolution, giving important insights into this signaling complex. Here we describe the biochemical protocol to study the interaction between GPCRs and G proteins before preparation of GPCR–G protein complexes for structural studies. We use gel filtration to analyze the binding properties between GPCR and G protein with the presence of agonist or antagonist, as well as the complex dissociation in the presence of GTP analogue. Methods used in the protocol are affinity purification and gel filtration, which are also commonly used in protein sample preparation for structural work. Therefore, the protocol can be easily adapted for large-scale sample preparation.

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Authors and Affiliations

  1. Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

    Filip Pamula & Ching-Ju Tsai

Authors
  1. Filip Pamula
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  2. Ching-Ju Tsai
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Correspondence to Ching-Ju Tsai .

Editor information

Editors and Affiliations

  1. School of Biological Sciences and School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, GA, USA

    Ingeborg Schmidt-Krey

  2. School of Physics, Georgia Institute of Technology, Atlanta, GA, USA

    James C. Gumbart

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Pamula, F., Tsai, CJ. (2021). Biochemical Characterization of GPCR–G Protein Complex Formation. In: Schmidt-Krey, I., Gumbart, J.C. (eds) Structure and Function of Membrane Proteins. Methods in Molecular Biology, vol 2302. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1394-8_3

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  • DOI: https://doi.org/10.1007/978-1-0716-1394-8_3

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1393-1

  • Online ISBN: 978-1-0716-1394-8

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