Antagonist binding profile of the split chimeric muscarinic m2-trunc/m3-tail receptor
ABSTRACT Recent evidence suggests that G-protein-coupled receptors can behave as multiple subunit receptors, and can be split into parts, maintaining their binding ability. Transfection of a truncated muscarinic m2 receptor (containing transmembrane domains I-V, named m2-trunc) with a gene fragment coding for the carboxyl-terminal receptor portion of the muscarinic m3 receptor (containing transmembrane domains VI and VII, named m3-tail) results in the formation of a binding site with a high affinity for the muscarinic ligand N-[3H]methylscopolamine. In this paper we analyse the antagonist binding profile of this chimeric m2-trunc/m3-tail receptor in comparison with the wild-type muscarinic m2 and m3 receptors. While many of the substances tested had an intermediate affinity for the chimeric m2-trunc/m3-tail receptor compared with m2 and m3, some compounds were able to distinguish between the chimeric m2-trunc/m3-tail receptor on the one hand and the m2 or the m3 receptor on the other. Among them, tripitramine (a high-affinity M2 receptor antagonist) bound to the m2-trunc/m3-tail receptor with the same affinity as m2, but it bound to the m3 receptor with a 103-fold lower affinity; pirenzepine (a selective muscarinic M1 receptor antagonist) bound to the chimeric receptor with an affinity that was 12- and 3-fold higher than that of m2 and m3, respectively. The results of this study demonstrate that the chimeric m2-trunc/m3-tail receptor has a pharmacological profile distinct from that of the originating muscarinic m2 and m3 receptors.
- SourceAvailable from: Graeme Milligan
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- "Transmembrane helices I-V and VI-VII form essentially independent folding domains. GPCR sequences can be split between transmembrane helices V and VI, and, when these fragments are co-expressed, functional receptors can be reconstituted (Maggio et al., 1996; Barbier et al., 1998; Scarselli et al., 2000). Co-expression of a reciprocal chimaera containing transmembrane regions I-V of the α2C adrenoceptor and helices VI and VII of the muscarinic m3 acetylcholine receptor, or vice versa, results in the reconstitution of binding sites for ligands for both of the native receptors. "
ABSTRACT: A range of approaches have recently provided evidence that G-protein-coupled receptors can exist as oligomeric complexes. Both homo-oligomers, comprising multiple copies of the same gene product, and hetero-oligomers containing more than one receptor have been detected. In several, but not all, examples, the extent of oligomerisation is regulated by the presence of agonist ligands, and emerging evidence indicates that receptor hetero-oligomers can display distinct pharmacological characteristics. A chaperonin-like role for receptor oligomerisation in effective delivery of newly synthesised receptors to the cell surface is a developing concept, and recent studies have employed a series of energy-transfer techniques to explore the presence and regulation of receptor oligomerisation in living cells. However, the majority of studies have relied largely on co-immunoprecipitation techniques, and there is still little direct information on the fraction of receptors existing as oligomers in intact cells.Journal of Cell Science 05/2001; 114(Pt 7):1265-71. · 5.33 Impact Factor
- Annual reports in medicinal chemistry 01/2000; 35:271-279. DOI:10.1016/S0065-7743(00)35025-4 · 1.19 Impact Factor