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G Protein Signaling

Methods and Protocols

  • Book
  • © 2004

Overview

Editors:
  1. Alan V. Smrcka

    1. Department of Pharmacology and Physiology, University of Rochester School of Medicine, Rochester

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Part of the book series: Methods in Molecular Biology (MIMB, volume 237)

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Table of contents (20 protocols)

  1. Front Matter

    Pages i-xii
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  2. Purification of G Protein Subunits and G Protein Effectors

    1. Front Matter

      Pages 1-1
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    2. Purification of Recombinant G Protein α Subunits from Escherichia coli

      • Wendy K. Greentree, Maurine E. Linder
      Pages 3-20

    3. Purification of G Protein Subunits from Sf9 Insect Cells Using Hexahistidine-Tagged α and βγ Subunits

      • Tohru Kozasa
      Pages 21-38

    4. Expression and Purification of Soluble Adenylyl Cyclase from Escherichia coli

      • Jeff A. Beeler, Wei-Jen Tang
      Pages 39-53

    5. Purification of Phospholipase C β and Phospholipase C ε from Sf9 Cells

      • Mousumi Ghosh, Huan Wang, Grant G. Kelley, Alan V. Smrcka
      Pages 55-64

  3. Assays for G Protein—Dependent Effector Activation In Vitro

    1. Front Matter

      Pages 65-65
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    2. Assay for G Protein-Dependent Activation of Phospholipase C β Using Purified Protein Components

      • Mousumi Ghosh, Alan V. Smrcka
      Pages 67-75

    3. Assays of Recombinant Adenylyl Cyclases Expressed in Sf9 Cells

      • Ronald Taussig
      Pages 77-85

  4. Assays For G Protein-Coupled Receptor-Mediated and G Protein Subunit-Mediated Effector Activation in Intact Cells

    1. Front Matter

      Pages 87-87
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    2. Measurement of G Protein-Coupled Receptor-Stimulated Phospholipase D Activity in Intact Cells

      • Stephanie J. Walker, H. Alex Brown
      Pages 89-97

    3. Analysis of G Protein-Mediated Activation of Phospholipase C in Cultured Cells

      • Bo Liu, Dianqing Wu
      Pages 99-102

  5. Receptor Mechanisms, Activity, and Localization

    1. Front Matter

      Pages 103-103
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    2. Intensive Mutational Analysis of G Protein-Coupled Receptors in Yeast

      • Andjelka Ćelić, Sara M. Connelly, Negin P. Martin, Mark E. Dumont
      Pages 105-120

    3. Green Fluorescent Protein-Tagged β-Arrestin Translocation as a Measure of G Protein-Coupled Receptor Activation

      • Stephen S. G. Ferguson, Marc G. Caron
      Pages 121-126

    4. Detection of G Protein-Coupled Receptors by Immunofluorescence Microscopy

      • Patricia M. Hinkle, John A. Puskas
      Pages 127-134

    5. Assay of G Protein-Coupled Receptor Activation of G Proteins in Native Cell Membranes Using [35S]GTPγS Binding

      • Jean M. Bidlack, Amy L. Parkhill
      Pages 135-143

    6. Analysis of the Coupling of G12/13 to G Protein-Coupled Receptors Using a Luciferase Reporter Assay

      • Bo Liu, Dianqing Wu
      Pages 145-149

  6. Disruption of Endogenous G Protein-Signaling Systems to Understand the Role of G Protein Subunits in Intact Systems

    1. Front Matter

      Pages 151-151
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    2. Selective Inhibition of G Protein-Mediated Pathways Using RGS Domains

      • Tohru Kozasa, Richard D. Ye
      Pages 153-167
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About this book

G proteins and G protein-coupled receptors are ubiquitously expressed proteins that regulate a wide range of physiological processes and are consequently the targets of many pharmaceuticals. In G Protein Signaling: Methods and Protocols, leading researchers describe in detail key methods for investigating G protein signaling from a variety of perspectives ranging from in vitro biochemistry to whole animal studies. The authors focus on the mechanisms of G protein and G protein-coupled receptor function and the roles of G protein subunits in cell biology and disease. Among the readily reproducible techniques presented are those for the purification of G proteins and effector enzymes, assays of these purified G proteins and effector enzymes, and for the study of G protein interactions with effectors in intact cells. Additional methods are provided for assaying G protein-coupled receptor structure, function, and localization, for studying the physiological roles for endogenous G proteins, and for examining lipid and phosphate modifications of RGS proteins. Each fully tested protocol includes a background introduction explaining the principle behind the technique, equipment and reagent lists, tips on troubleshooting and avoiding known pitfalls, and, where needed, a discussion of the interpretation and use of results.
Comprehensive and highly practical, G Protein Signaling: Methods and Protocols offers novice and expert researchers alike an array of cutting-edge tools to illuminate the roles of G protein and G protein-coupled receptors, as well as a better understanding of their action with drugs, neurotransmitters, and sensory stimuli.

Reviews

"...a timely and useful set of protocols...a significant contribution." -Doody's Health Sciences Book Review Journal

Editors and Affiliations

  • Department of Pharmacology and Physiology, University of Rochester School of Medicine, Rochester

    Alan V. Smrcka

Bibliographic Information

  • Book Title: G Protein Signaling

  • Book Subtitle: Methods and Protocols

  • Editors: Alan V. Smrcka

  • Series Title: Methods in Molecular Biology

  • DOI: https://doi.org/10.1385/1592594301

  • Publisher: Humana Totowa, NJ

  • eBook Packages: Springer Protocols

  • Copyright Information: Humana Press 2004

  • Hardcover ISBN: 978-1-58829-137-0Published: 15 September 2003

  • Softcover ISBN: 978-1-61737-381-7Published: 09 December 2010

  • eBook ISBN: 978-1-59259-430-6Published: 02 February 2008

  • Series ISSN: 1064-3745

  • Series E-ISSN: 1940-6029

  • Edition Number: 1

  • Number of Pages: XII, 250

  • Topics: Cell Biology

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