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G 16/z Chimeras Efficiently Link a Wide Range of G Protein— Coupled Receptors to Calcium Mobilization

Department of Biochemistry, the Molecular Neuroscience Center, and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China

Maurice K.C. Ho

Department of Biochemistry, the Molecular Neuroscience Center, and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China

Cecilia S.S. Wong

Department of Biochemistry, the Molecular Neuroscience Center, and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China

Jasmine H.P. Chan

Department of Biochemistry, the Molecular Neuroscience Center, and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China

Anson H.M. Pau

Department of Biochemistry, the Molecular Neuroscience Center, and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China

Yung H. Wong

Department of Biochemistry, the Molecular Neuroscience Center, and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China, boyung{at}ust.hk

G protein—coupled receptors (GPCRs) represent a class of important therapeutic targets for drug discovery. The integration of GPCRs into contemporary high-throughput functional assays is critically dependent on the presence of appropriate G proteins. Given that different GPCRs can discriminate against distinct G proteins, a universal G protein adapter is extremely desirable. In this report, the authors evaluated two highly promiscuous G_16/z chimeras, 16z25 and 16z44, for their ability to translate GPCR activation into Ca²⁺ mobilization using the fluorescence imaging plate reader (FLIPR) and aequorin. A panel of 24 G_s- or G_i-coupled receptors was examined for their functional association with the G_16/z chimeras. Although most of the GPCRs tested were incapable of inducing Ca²⁺ mobilization upon their activation by specific agonists, the introduction of 16z25 or 16z44 allowed all of these GPCRs to mediate agonist-induced Ca ²⁺ mobilization. In contrast, only 16 of the GPCRs tested were capable of using G₁₆ to mobilize intracellular Ca²⁺. Analysis of dose-response curves obtained with the -opioid, dopamine D₁ , and Xenopus melatonin Mel1c receptors revealed that the G _16/z chimeras possess better sensitivity than G₁₆ in both the FLIPR and aequorin assays. Collectively, these studies help to validate the promiscuity of the G_16/z chimeras as well as their application in contemporary drug-screening assays that are based on ligand-induced Ca ²⁺ mobilization. (Journal of Biomolecular Screening 2003:39-49)

Key Words: G proteins • GPCR • Ca2+ mobilization • aequorin • G16

Journal of Biomolecular Screening, Vol. 8, No. 1, 39-49 (2003)
DOI: 10.1177/1087057102239665


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