Hishinuma Shigeru, Kosaka Kiyoe, Akatsu Chizuru, Uesawa Yoshihiro, Fukui Hiroyuki, Shoji Masaru
Department of Pharmacodynamics, Meiji Pharmaceutical University, 2-522-1 Noshio, Kiyose, Tokyo 204-8588, Japan.
Department of Pharmacodynamics, Meiji Pharmaceutical University, 2-522-1 Noshio, Kiyose, Tokyo 204-8588, Japan.
Biochem Pharmacol. 2017 Mar 15;128:46-54. doi: 10.1016/j.bcp.2016.12.021. Epub 2016 Dec 29.
The affinity of ligands for G-protein-coupled receptors (GPCRs) is allosterically regulated by Na via a highly conserved aspartate residue (Asp) in the second transmembrane domain of GPCRs. In the present study, we examined the Na-mediated regulation of the affinity of ligands for G-protein-coupled human histamine H receptors in Chinese hamster ovary cells. The affinities of 3 agonists and 20 antihistamines were evaluated by their displacement curves against the binding of [H]-mepyramine to membrane preparations in the presence or absence of 100mM NaCl. The affinities of most drugs including histamine, an agonist, and d-chlorpheniramine, a first-generation antihistamine, were reduced by NaCl, with the extent of NaCl-mediated changes varying widely between drugs. In contrast, the affinities of some second-generation antihistamines such as fexofenadine were increased by NaCl. These changes were retained in intact cells. The mutation of Asp (Asp73) to asparagine abrogated NaCl-induced reductions in affinities for histamine and d-chlorpheniramine, but not NaCl-induced increases in the affinity for fexofenadine. Quantitative structure-activity relationship (QSAR) analyses showed that these Na-mediated changes were explained and predicted by a combination of the molecular energies and implicit solvation energies of the compounds. These results suggest that Na diversely regulates the affinity of ligands for H receptors from the extracellular sites of receptors via Asp73-dependent and -independent mechanisms in a manner that depends on the physicochemical properties of ligands. These results may contribute to a deeper understanding of the fundamental mechanisms by which the affinity of ligands for their receptors is allosterically regulated by Na.
配体与G蛋白偶联受体（GPCRs）的亲和力通过GPCRs第二个跨膜结构域中一个高度保守的天冬氨酸残基（Asp）由Na进行变构调节。在本研究中，我们检测了Na对中国仓鼠卵巢细胞中G蛋白偶联的人组胺H受体配体亲和力的调节作用。通过在存在或不存在100mM NaCl的情况下，3种激动剂和20种抗组胺药针对[H]-美吡拉敏与膜制剂结合的置换曲线来评估它们的亲和力。包括激动剂组胺和第一代抗组胺药d-氯苯那敏在内的大多数药物的亲和力被NaCl降低，不同药物之间NaCl介导的变化程度差异很大。相反，一些第二代抗组胺药如非索非那定的亲和力被NaCl增加。这些变化在完整细胞中得以保留。将Asp（Asp73）突变为天冬酰胺消除了NaCl诱导的组胺和d-氯苯那敏亲和力降低，但没有消除NaCl诱导的非索非那定亲和力增加。定量构效关系（QSAR）分析表明，这些Na介导的变化可以通过化合物的分子能量和隐式溶剂化能量的组合来解释和预测。这些结果表明，Na通过依赖于Asp73和不依赖于Asp73的机制，从受体的细胞外位点以取决于配体物理化学性质的方式，对H受体配体的亲和力进行不同的调节。这些结果可能有助于更深入地理解Na变构调节配体与其受体亲和力的基本机制。
