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Expanding the Concept of G Protein-Coupled Receptor (GPCR) Dimer Asymmetry towards GPCR-Interacting Proteins
Pharmaceuticals 2011, 4(3), 457-469; doi:10.3390/ph4030457

Dual-Color Bioluminescence Analysis for Quantitatively Monitoring G-Protein-Coupled Receptor and β-Arrestin Interactions

1, 1, 1 and 1,2,*
1 Department of Chemistry, School of Science, The University of Tokyo, Bunkyo-ku, Hongo, Tokyo 113, Japan 2 PRESTO, Japan Science and Technology Agency, Tokyo, Japan
* Author to whom correspondence should be addressed.
Received: 13 December 2010 / Revised: 17 February 2011 / Accepted: 18 February 2011 / Published: 25 February 2011
(This article belongs to the Special Issue GPCR Based Drug Discovery)
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G protein-coupled receptors (GPCRs) are crucial elements in mammalian signal transduction, and are considered to represent potent drug targets. We have previously developed a GPCR assay system in cultured cells based on complementation of split fragments of click beetle (Pyrearinus termitilluminans) luciferase. The interaction of GPCRs with its target, β-arrestin, resulted in strong emission of bioluminescence upon stimulation with its specific ligand. In this study, we improved precision of the GPCR assay system by using railroad worm (Phrixothrix hirtus) luciferase as an internal control. We generated stable cell lines harboring the railroad worm luciferase and quantitatively evaluate the extent of GPCR-β-arrestin interactions. We showed concentration-dependent bioluminescence responses for four GPCRs: β2-adrenoceptor, endothelin receptor type A, α2-adrenoceptor and human μ-opioid receptor. We also demonstrated that the variation of responses was reduced significantly by normalizing the data with bioluminescence from railroad worm luciferase. This assay system represents a simple and reliable approach for screening drug candidates in a high throughput manner.
Keywords: GPCR; luciferase; protein interaction GPCR; luciferase; protein interaction
This is an open access article distributed under the Creative Commons Attribution License (CC BY) which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Kafi, A.; Hattori, M.; Misawa, N.; Ozawa, T. Dual-Color Bioluminescence Analysis for Quantitatively Monitoring G-Protein-Coupled Receptor and β-Arrestin Interactions. Pharmaceuticals 2011, 4, 457-469.

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