Woo Anthony Yiu-Ho, Wang Tian-Bing, Zeng Xiaokun, Zhu Weizhong, Abernethy Darrell R, Wainer Irving W, Xiao Rui-Ping
Laboratory of Cardiovascular Science, Gerontology Research Center, NIA, National Institutes of Health, 5600 Nathan Shock Drive, Baltimore, MD 21224, USA.
Mol Pharmacol. 2009 Jan;75(1):158-65. doi: 10.1124/mol.108.051078. Epub 2008 Oct 7.
A fundamental question regarding receptor-G protein interaction is whether different agonists can lead a receptor to different intracellular signaling pathways. Our previous studies have demonstrated that although most beta(2)-adrenoceptor agonists activate both G(s) and G(i) proteins, fenoterol, a full agonist of beta(2)-adrenoceptor, selectively activates G(s) protein. Fenoterol contains two chiral centers and may exist as four stereoisomers. We have synthesized a series of stereoisomers of fenoterol and its derivatives and characterized their receptor binding and pharmacological properties. We tested the hypothesis that the stereochemistry of an agonist determines selectivity of receptor coupling to different G protein(s). We found that the R,R isomers of fenoterol and methoxyfenoterol exhibited more potent effects to increase cardiomyocyte contraction than their S,R isomers. It is noteworthy that although (R,R)-fenoterol and (R,R)-methoxyfenoterol preferentially activate G(s) signaling, their S,R isomers were able to activate both G(s) and G(i) proteins as evidenced by the robust pertussis toxin sensitivities of their effects on cardiomyocyte contraction and on phosphorylation of extracellular signal-regulated kinase 1/2. The differential G protein selectivities of the fenoterol stereoisomers were further confirmed by photoaffinity labeling studies on G(s),G(i2), and G(i3) proteins. The inefficient G(i) signaling with the R,R isomers is not caused by the inability of the R,R isomers to trigger the protein kinase A (PKA)-mediated phosphorylation of the beta(2)-adrenoceptor, because the R,R isomers also markedly increased phosphorylation of the receptor at serine 262 by PKA. We conclude that in addition to receptor subtype and phosphorylation status, the stereochemistry of a given agonist plays an important role in determining receptor-G protein selectivity and downstream signaling events.
一个关于受体 - G蛋白相互作用的基本问题是，不同的激动剂是否能使受体引发不同的细胞内信号通路。我们之前的研究表明，尽管大多数β₂ - 肾上腺素能受体激动剂能同时激活Gₛ和Gᵢ蛋白，但β₂ - 肾上腺素能受体的完全激动剂非诺特罗却选择性地激活Gₛ蛋白。非诺特罗含有两个手性中心，可能以四种立体异构体的形式存在。我们合成了一系列非诺特罗及其衍生物的立体异构体，并对它们的受体结合和药理特性进行了表征。我们检验了这样一个假设，即激动剂的立体化学决定受体与不同G蛋白偶联的选择性。我们发现，非诺特罗和甲氧基非诺特罗的R,R异构体在增强心肌细胞收缩方面比它们的S,R异构体表现出更强的作用。值得注意的是，尽管(R,R)-非诺特罗和(R,R)-甲氧基非诺特罗优先激活Gₛ信号，但它们的S,R异构体能够同时激活Gₛ和Gᵢ蛋白，这一点从它们对心肌细胞收缩和细胞外信号调节激酶1/2磷酸化作用的强烈百日咳毒素敏感性中得到证明。非诺特罗立体异构体不同的G蛋白选择性通过对Gₛ、Gᵢ₂和Gᵢ₃蛋白的光亲和标记研究得到了进一步证实。R,R异构体Gᵢ信号传导效率低下并非是由于R,R异构体无法触发蛋白激酶A（PKA）介导的β₂ - 肾上腺素能受体磷酸化，因为R,R异构体也显著增加了PKA介导的受体丝氨酸262位点的磷酸化。我们得出结论，除了受体亚型和磷酸化状态外，特定激动剂的立体化学在决定受体 - G蛋白选择性和下游信号事件中起着重要作用。
