Pharmaceuticals 2011, 4(3), 457-469; doi:10.3390/ph4030457
Article

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

A.K.M. Kafi 1 , Mitsuru Hattori 1 , Naomi Misawa 1  and Takeaki Ozawa 1,2,* email
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; in revised form: 17 February 2011 / Accepted: 18 February 2011 / Published: 25 February 2011
(This article belongs to the Special Issue GPCR Based Drug Discovery)
PDF Full-text Download PDF Full-Text [1028 KB, uploaded 25 February 2011 11:49 CET]
Abstract: 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

Article Statistics

Click here to load and display the download statistics.

Cite This Article

MDPI and ACS Style

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.

AMA Style

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(3):457-469.

Chicago/Turabian Style

Kafi, A.K.M.; Hattori, Mitsuru; Misawa, Naomi; Ozawa, Takeaki. 2011. "Dual-Color Bioluminescence Analysis for Quantitatively Monitoring G-Protein-Coupled Receptor and β-Arrestin Interactions." Pharmaceuticals 4, no. 3: 457-469.

Pharmaceuticals EISSN 1424-8247 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert