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Characterization of G Protein-coupled Receptors by a Fluorescence-based Calcium Mobilization Assay
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Measuring GPCR-Induced Intracellular Calcium Signaling Using a Quantitative High-Throughput Assay

Shree S Kumar1, Kathleen G Mountjoy2

  • 1Faculty of Medical and Health Sciences, Department of Molecular Medicine and Pathology, School of Medical Sciences, The University of Auckland, Auckland, New Zealand.

Methods in Molecular Biology (Clifton, N.J.)
|October 12, 2024
PubMed

Keywords:
Calcium mobilizationFluorescenceFura-2 dyeG-protein-coupled receptorsIntracellular calcium quantitationPHERAstar FS plate readerRatiometric

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G Protein-coupled Receptors

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G protein-coupled receptor (GPCR) signaling plays a crucial role in cell functioning. GPCR desensitization is an equally essential process. It allows cells to respond to changing environments and regain sensitivity to new stimuli while preventing unnecessary stimulation when no longer needed. Prolonged exposure to stimuli leads to GPCR desensitization. It involves blocking the receptors from binding and activating additional G proteins. This inhibits activation of downstream effectors, thereby...
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GPCRs Regulate Adenylyl Cylase Activity

Some GPCRs transmit signals through adenylyl cyclase (AC), a transmembrane enzyme. AC helps synthesize second messenger cyclic adenosine monophosphate (cAMP). AC catalyzes cyclization reaction and converts ATP to cAMP by releasing a pyrophosphate. The pyrophosphate is further hydrolyzed to phosphate by the enzyme pyrophosphatase, which drives cAMP synthesis to completion. However, cAMP is rapidly degraded to 5′ AMP by the enzymes phosphodiesterase (PDE), preventing overstimulation of cells.
Two...
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G Protein–Coupled Receptors (GPCRs) are membrane-bound receptors that transiently associate with heterotrimeric G proteins and induce an appropriate response to various stimuli. GPCRs regulate critical physiological pathways and are excellent drug targets for treating diseases such as diabetes, cancer, obesity, depression, or Alzheimer's. Nearly 35% of approved drugs implement their therapeutic effects by selectively interacting with specific GPCRs.
GPCRs are also called heptahelical, 7TM, or...