Joshi S, Lee J W, Wong Y H
Department of Biology and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Kowloon, China.
Eur J Neurosci. 1999 Feb;11(2):383-8. doi: 10.1046/j.1460-9568.1999.00442.x.
Opioid receptors (mu, delta and kappa) are known to regulate diverse physiological functions and yet, at the molecular level, they are coupled to a seemingly identical set of G proteins. A recent study has discerned subtle differences between the opioid receptors in their ability to activate the pertussis toxin-insensitive G16. Differences in microarchitecture might be magnified when these receptors are provided with 'non-native' partners. Here, we examined whether the opioid receptors can interact productively with a set of chimeric Galphaq subunits which are known to link many Gi-coupled receptors to phosphoinositide-specific phospholipase C (PI-PLC). The qi5, qo5 and qz5 chimeras have the last five residues of Galphaq replaced by those of Galphai, Galphao and Galphaz, respectively. Except for mu-receptor and qo5, each pair of opioid receptor and Galphaq chimera allowed opioid agonists to stimulate PI-PLC in transfected COS-7 cells. The Galphaq chimera-mediated responses were ligand selective, agonist dose dependent and saturable. The most robust responses were obtained with kappa-receptor and qi5 or qz5, whereas the coupling of delta- and mu-receptors to Galphaq chimeras produced much weaker responses. Among the Galphaq chimeras, qo5 was less efficiently coupled to the opioid receptors. As revealed by radioligand binding assays and immunoblot analysis, differences in the efficiency of coupling were not due to variations in the expression of receptors and Galphaq chimeras. Differences in the magnitude of PI-PLC responses are thus likely to represent structural incompatibility between opioid receptors and Galphaq chimeras, suggesting that each opioid receptor possesses unique structural surfaces for the binding of G proteins.
已知阿片受体（μ、δ和κ）可调节多种生理功能，但在分子水平上，它们与一组看似相同的G蛋白偶联。最近的一项研究发现，阿片受体在激活百日咳毒素不敏感的G16的能力上存在细微差异。当这些受体与“非天然”伴侣结合时，微观结构上的差异可能会被放大。在这里，我们研究了阿片受体是否能与一组嵌合Gαq亚基有效相互作用，已知这些亚基可将许多与Gi偶联的受体与磷酸肌醇特异性磷脂酶C（PI-PLC）连接起来。qi5、qo5和qz5嵌合体的Gαq的最后五个残基分别被Gαi、Gαo和Gαz的相应残基取代。除了μ受体和qo5外，每一对阿片受体和Gαq嵌合体都能使阿片激动剂在转染的COS-7细胞中刺激PI-PLC。Gαq嵌合体介导的反应具有配体选择性、激动剂剂量依赖性和饱和性。κ受体与qi5或qz5结合时产生的反应最强烈，而δ和μ受体与Gαq嵌合体的偶联产生的反应则弱得多。在Gαq嵌合体中，qo5与阿片受体的偶联效率较低。放射性配体结合试验和免疫印迹分析表明，偶联效率的差异不是由于受体和Gαq嵌合体表达的变化所致。因此，PI-PLC反应幅度的差异可能代表阿片受体与Gαq嵌合体之间的结构不兼容性，这表明每个阿片受体都具有独特的G蛋白结合结构表面。
