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Defining the Functional Equivalence of Wild-Type and Chemically Engineered G Protein-Coupled Receptors

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Book cover Designer Receptors Exclusively Activated by Designer Drugs

Part of the book series: Neuromethods ((NM,volume 108))

Abstract

The functional connection between activation of a single G protein-coupled receptor (GPCR) and a specific physiological outcome may be obscured by the presence of other closely related members of the GPCR superfamily that share spatiotemporal patterns of expression and equipotent activation by endogenous ligands. To address this issue, molecular and chemical genetic techniques have been developed to generate mutationally modified GPCRs only susceptible to activation by one or more synthetic ligands that are inactive at the equivalent wild-type receptor. This chapter provides both an overview of strategies used to generate such “receptors activated solely by synthetic ligands” (RASSLs) and, in more detail, approaches that have been used to assess the equivalence or otherwise of function of a wild-type and corresponding RASSL receptor. The human muscarinic M3 acetylcholine receptor is used as the exemplar but similar preliminary studies should be employed before further use, particularly in vivo, of such RASSLs.

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Abbreviations

BAL:

2,3-Dimercapto-1-propanol

BRET:

Bioluminescence resonance energy transfer

CFP:

Cyan fluorescent protein

CNO:

Clozapine N-oxide

DREADD:

Designer receptor exclusively activated by designer drugs

EDT:

2-Ethanedithiol

FRET:

Fluorescence resonance energy transfer

Fura-2AM:

Fura-2 acetoxymethyl ester

GFP:

Green fluorescent protein

GPCR:

G protein-coupled receptor

IL3:

Third intracellular loop

IP1:

Inositol monophosphate

RASSL:

Receptor activated solely by synthetic ligands

TMD:

Transmembrane domain

YFP:

Yellow fluorescent protein

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Authors and Affiliations

  1. Molecular Pharmacology Group, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, University Avenue, Glasgow, G12 8QQ, UK

    Elisa Alvarez-Curto & Graeme Milligan

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  1. Elisa Alvarez-Curto
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  2. Graeme Milligan
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Correspondence to Graeme Milligan .

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Editors and Affiliations

  1. Department of Medical Biochemistry and Molecular Biology, University of Saarland Medical Faculty, Homburg, Germany

    Gerald Thiel

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Alvarez-Curto, E., Milligan, G. (2015). Defining the Functional Equivalence of Wild-Type and Chemically Engineered G Protein-Coupled Receptors. In: Thiel, G. (eds) Designer Receptors Exclusively Activated by Designer Drugs. Neuromethods, vol 108. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2944-3_1

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  • DOI: https://doi.org/10.1007/978-1-4939-2944-3_1

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