Functional coupling of the human dopamine D2 receptor with Gαi1, Gαi2, Gαi3 and Gαo G proteins:: evidence for agonist regulation of G protein selectivity
Functional coupling of the human dopamine D2 receptor with Gαi1, Gαi2, Gαi3 and Gαo G proteins:: evidence for agonist regulation of G protein selectivity
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DOI：
10.1038/sj.bjp.0705116
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发表时间：
2003-03-01
影响因子：
7.3
通讯作者：
Strange, PG
中科院分区：
文献类型：
--
作者：
Gazi, L;Nickolls, SA;Strange, PG
1利用杆状病毒表达系统，在Sf9细胞中用四种不同的G蛋白表达人多巴胺D-2long （D-2L）受体。当与G(i)/G(o) G蛋白(G(i1) α， G(i2) α， G(i3) α或G(o) α，加上Gbeta（1)和Ggamma(2)）共表达时，受体显示出对激动剂（多巴胺和NPA）的高亲和力结合位点，对GTP （100 mum）敏感，表明受体与不同的G蛋白之间存在相互作用采用[H-3]-spiperone饱和结合(R)和[S-35]-GTPgammaS饱和结合(G)评价受体与G蛋白的比值（R: G ratio）。G（i1）、G（i2）、G（i3）和Go的R: G比值分别为1:12、1:3、1:14和1:5。然而，当G（i2）和G(o)的R:G比为1:2和1:12时，对[S-35]-GTPgammaS结合的刺激没有发现差异我们测试了几种激动剂刺激[S-35]-GTPgammaS结合到共表达受体和各种G蛋白的膜上的能力。所有化合物在表达G（i3）和G(o)的制剂中均表现出激动剂活性。然而，对于G（i2）和G（i1）制剂，S-(-)-3- ppp和对酪胺等化合物无法刺激[S-35]- gtpys结合大多数化合物在表达G(o)的制剂中表现出更高的相对功效（与多巴胺相比）和更高的效力。不同激动剂在不同制剂中的作用比较表明，每种激动剂对4种G蛋白的激活程度不同我们得出结论，G蛋白被D-2L受体激活的选择性程度取决于受体的构象。
1 The human dopamine D-2long (D-2L) receptor was expressed with four different G proteins in Sf9 cells using the baculovirus expression system. When co-expressed with G(i)/G(o) G proteins (G(i1)alpha, G(i2)alpha, G(i3)alpha, or G(o)alpha, plus Gbeta(1) and Ggamma(2)) the receptor displayed a high-affinity binding site for the agonists (dopamine and NPA), which was sensitive to GTP (100 mum), demonstrating interaction between the receptor and the different G proteins.2 The receptor to G protein ratio (R: G ratio) was evaluated using [H-3]-spiperone saturation binding (R) and [S-35]-GTPgammaS saturation binding (G). R: G ratios of 1: 12, 1: 3, 1: 14 and 1: 5 were found for G(i1), G(i2), G(i3), and Go preparations, respectively. However, when R:G ratios of 1:2 and 1: 12 were compared for G(i2) and G(o), no difference was found for the stimulation of [S-35]-GTPgammaS binding.3 Several agonists were tested for their ability to stimulate [S-35]-GTPgammaS binding to membranes co-expressing the receptor and various G proteins. All the compounds tested showed agonist activity in preparations expressing G(i3) and G(o). However, for G(i2) and G(i1) preparations, compounds such as S-(-)-3-PPP and p-tyramine were unable to stimulate [S-35]-GTPyS binding.4 Most of the compounds showed higher relative efficacies (compared to dopamine) and higher potencies in the preparation expressing G(o). Comparison of the effects of different agonists in the different preparations showed that each agonist differentially activates the four G proteins.5 We conclude that the degree of selectivity of G protein activation by the D-2L receptor can depend on the conformation of the receptor stabilised by an agonist.