G Protein Signaling : Methods and Protocols

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.

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.