To explore the molecular basis of ligand binding and receptor activation of human motilin receptor

背景。胃动素（Motilin）和胃饥饿素（ghrelin）被认为是哺乳动物胃肠运动功能的重要内源性调节剂，分别由胃动素受体和密切相关的生长激素分泌素（GHS）受体介导。本研究的目的是确定人胃动素受体内的配体结合位点，探索胃动素和GHS受体在人胃肠道中的分布，并确定人胃动素受体的分子性质。方法。我们使用受体诱变和光亲和标记的方法检测了人胃动素受体的细胞外结构域。我们还分析了人胃肠道中胃动素和GHS受体的mRNA表达水平和免疫反应性。结果。运动素受体Cys25、Cys30、Cys111和Cys235的氨基端和第一、第二胞外环结构域的胱氨酸残基对于肽配体（运动素）和非肽基配体（红霉素）的生物活性以及保存该受体中的二硫键至关重要。膜周残基Gly36、Pro103、Leu109和Phe332仅对肽配体的结合和生物活性有重要作用。长形式的胃动素受体，而不是短形式的，在胃肠道的所有部位都有表达，并且在肌肉中的表达水平高于粘膜。胃动素受体免疫反应性存在于肌肉细胞和肌肠丛。GHS受体mRNA和免疫反应性在胃肠道各部位粘膜层和肌肉层表达相同。结论。这些结果表明，胃动素受体对肽配体和非肽配体结合和作用的细胞外结构域的不同贡献，以及胃动素受体沿整个胃肠道的弥漫性肌肉表达，使其成为胃动素类药物治疗运动障碍的有用潜在靶点。

Background. Motilin and ghrelin have been recognized as important endogenous regulators of gastrointestinal (GI) motor function in mammals, mediated by the motilin receptor and by the closely-related growth hormone secretagogue (GHS) receptor, respectively. The aims of this study were to identify the ligand-binding sites within the human motilin receptor, and to explore the distribution of motilin and GHS receptors along the human gastrointestinal tract, and to establish the molecular nature of the human motilin receptor. Methods. We examined the extracellular domains of the human motilin receptor using the methods of receptor mutagenesis and photoaffinity labeling. We also analyzed levels of expression of mRNA and immunoreactivity for motilin and GHS receptors in the human GI tract. Results. Cys residues in the amino terminus and the first and second extracellular loop domains in the motilin receptor, Cys25, Cys30, Cys111, and Cys235, are critical for both peptide ligand, motilin, and non-peptidyl ligand, erythromycin, biological activities, conserving the disulfide bond in this receptor. The perimembranous residues, Gly36, Pro103, Leu109, and Phe332, are important only for binding and biological activity of peptide ligand. The long form of the motilin receptor, but not the short form, was expressed in all parts of the GI tract, and expressed at higher levels in muscle than in mucosa. Motilin receptor immunoreactivity was present on muscle cells and myenteric plexus. In contrast, GHS receptor mRNA and immunoreactivity were expressed equally in both mucosal and muscle layers in all parts of GI tract. Conclusions. These results, suggest that differential contribution of motilin receptor extracellular domains for peptide ligand and non-peptidyl ligand binding and action, and the diffuse muscle expression of the motilin receptor along the entire GI tract make it a useful potential target for motilide drugs for dysmotility.