G protein signaling : methods and protocols
著者
書誌事項
G protein signaling : methods and protocols
(Methods in molecular biology / John M. Walker, series editor, v. 237)
Humana Press, c2004
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注記
Includes bibliographical references and index
内容説明・目次
内容説明
Since the initial discovery of the G protein-coupled receptor system that regulates cyclicAMP production, the G protein field has rapidly expanded. Cell surface receptors that couple to heterotrimeric G proteins, the G prote- coupled receptors (GPCRs), number in the hundreds and bind to a wide div- sity of ligands including, biogenic amines (e. g. , adrenaline), lipid derivatives (e. g. , lysophosphatidic acid), peptides (e. g. , opioid peptides), proteins (e. g. , thyroid-stimulating hormone), and odorants to name a few. The GPCR system is found throughout biology in such simple organisms as yeast and in such more complex organisms as Dictyostelium discoideum (slime mold), Caen- habditis elegans (nematode worm), and of course in humans. GPCRs and their associated G protein systems are the subject of intense academic research and because of their involvement in a human biology and disease, the pharmac- tical industry has large research initiatives dedicated to the study of GPCRs. By some estimates, more than 50% of the pharmaceuticals on the market are targeted at GPCRs.
The G protein/G protein-coupled receptor system consists of a receptor (GPCR), a heterotrimeric G protein consisting of ?, ?, and ? subunits, and an effector. G protein effector molecules, such as enzymes or ion channels, respond to acti- tion by the G protein to generate second messengers or changes in membrane potential that lead to alterations in cell physiology.
目次
Part I. Purification of G Protein Subunits and G Protein Effectors
Purification of Recombinant G Protein a Subunits from Escherichia coli
Wendy K. Greentree and Maurine E. Linder
Purification of G Protein Subunits from Sf9 Insect Cells Using Hexahistidine-Tagged a and bg Subunits
Tohru Kozasa
Expression and Purification of Soluble Adenylyl Cyclase from Eschericia coli
Jeff A. Beeler and Wei-Jen Tang
Purification of Phospholipase C b and Phospholipase C e from Sf9 Cells
Mousumi Ghosh, Huan Wang, Grant G. Kelley, and Alan V. Smrcka
Part II. Assays for G Protein-Dependent Effector Activation In Vitro
Assay for G Protein-Dependent Activation of Phospholipase C b Using Purified Protein Components
Mousumi Ghosh and Alan V. Smrcka
Assays of Recombinant Adenylyl Cyclases Expressed in Sf9 Cells
Ronald Taussig
Part III. Assays for G Protein-Coupled Receptor-Mediated and G Protein Subunit-Mediated Effector Activation in Intact Cells
Measurement of G Protein-Coupled Receptor-Stimulated Phospholipase D Activity in Intact Cells
Stephanie J. Walker and H. Alex Brown
Analysis of G Protein-Mediated Activation of Phospholipase C in Cultured Cells
Bo Liu and Dianqing Wu
Part IV. Receptor Mechanisms, Activity, and Localization
Intensive Mutational Analysis of G Protein-Coupled Receptors in Yeast
Andjelka Celic, Sara M. Connelly, Negin P. Martin, and Mark E. Dumont
Green Fluorescent Protein-Tagged b-Arrestin Translocation as a Measure of G Protein-Coupled Receptor Activation
Stephen S. G. Ferguson and Marc G. Caron
Detection of G Protein-Coupled Receptors by Immunofluorescence Microscopy
Patricia M. Hinkle and John A. Puskas
Assay of G Protein-Coupled Receptor Activation of G Proteins in Native Cell Membranes Using [35S]GTPgS Binding
Jean M. Bidlack and Amy L.Parkhill
Analysis of the Coupling of G12/13 to G Protein-Coupled Receptors Using a Luciferase Reporter Assay
Bo Liu and Dianqing Wu
Part V. Disruption of Endogenous G Protein-Signaling Systems to Understand the Role of G Protein Subunits in Intact Systems
Selective Inhibition of G Protein-Mediated Pathways Using RGS Domains
Tohru Kozasa and Richard D. Ye
Ribozymes as Tools for Suppression of G Protein g Subunits
Janet D. Robishaw, Zheng-ping Guo, and Qin Wang
In Vivo Adenoviral-Mediated Gene Transfer of the bARKct to Study the Role of Gbg in Arterial Restenosis
Guido Iaccarino and Walter J. Koch
Part VI. Posttranslational Modifications of Regulators of G Protein Signaling
Analysis of RGS Protein Palmitoylation
Leah S. Bernstein, Maurine E. Linder, and John R. Hepler
Methods for Measuring RGS Protein Phosphorylation by G Protein-Regulated Kinases
Susanne Hollinger and John R. Hepler
Part VII. Use of GFP-Tagged G Proteins and Effectors in Intact Cells
The Use of Green Fluorescent Proteins to View Association Between Phospholipase Cb and G Protein Subunits in Cells
Suzanne Scarlata and Louisa Dowal
Cellular Localization of GFP-Tagged a Subunits
Thomas R. Hynes, Thomas E. Hughes, and Catherine H. Berlot
Index
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