Secretin Receptor Structure, Function and Regulation

描述（由申请人提供）：本申请的总体目标是更好地了解分泌素受体（一种原型B类G蛋白偶联受体）的配体诱导激活状态的分子基础。这些见解将填补当前知识的关键空白，并促进受体活性药物的最终开发。该受体家族包括治疗糖尿病、肥胖、骨质疏松、偏头痛、焦虑和抑郁的既定靶点；然而，开发作用于这些靶点的小分子激动剂尤其具有挑战性。最近解决的晶体结构展示了两个家族成员独特的开放螺旋束域，这有助于解释这一挑战。因此，该结构域已成为当前提案的焦点，其组分目标直接针对螺旋束与(i)生物活性配体，（ii）受体氨基端和（iii）相关膜蛋白相互作用的影响。中心假说认为，该结构域的构象变化引发了一系列细胞内信号转导事件，不同的配体直接与之相互作用，或通过与受体氨基末端的结合间接与之相互作用，以及与四元复合物内其他膜蛋白的相互作用，都会对其产生不同的影响。新认识到分泌素受体参与独特的跨类异受体复合物与血管紧张素1a受体提供了相关的生理学和治疗的见解，也有强有力的理由使用分泌素受体激动剂来管理相关的主要健康问题，包括肥胖，糖尿病，高血压，心力衰竭，以及胆汁淤滞。我们的具体目标将测试三个假设：(i)螺旋束高的开放空间，没有传统的对接口袋，在那里可以有效地映射正位激动剂的作用；（ii）氨基末端和核心结构域以特定和动态的方式相互作用，影响受体的激活状态；和

DESCRIPTION (provided by applicant): The overall objective of this application is to better understand the molecular basis of ligand-induced activation states of the secretin receptor, a prototypic class B G protein-coupled receptor. These insights should fill key gaps in current knowledge and facilitate the ultimate development of receptor-active drugs. This receptor family includes established targets for treatment of diabetes, obesity, osteoporosis, migraine, anxiety, and depression; however, development of small molecule agonists acting at these targets has been particularly challenging. Recently solved crystal structures demonstrating uniquely open helical bundle domains of two family members have helped to explain this challenge. This domain has, therefore, become the focal point of the current proposal, with component aims directed at the impact of helical bundle interactions with (i) biologically active ligands, (ii) th receptor amino terminus, and (iii) associated membrane proteins. The CENTRAL HYPOTHESIS is that conformational changes in this domain that initiate a spectrum of intracellular signaling events can be differentially affected by various ligands that interact with it directly, or indirecly by binding to the receptor amino terminus, and by interactions with other membrane proteins within quaternary complexes. New recognition of secretin receptor involvement in a unique cross-class hetero-receptor complex with the angiotensin 1a receptor provides insights relevant to physiology and therapeutics, and there is also strong rationale for use of a secretin receptor agonist to manage related major health problems, including obesity, diabetes, hypertension, and heart failure, as well as cholestasis. Our SPECIFIC AIMS will test three hypotheses: (i) the open space high in the helical bundle that is devoid of traditional docking pockets where the orthosteric agonist acts can be effectively mapped; (ii) amino-terminal and core domains interact in a specific and dynamic manner that affects the state of activation of the receptor; and (iii) key molecular interactions exist within a cross-class hetero-receptor complex that can be utilized to selectively modulate important biological activities. The first aim will explore the molecular determinants of agonist activity by utilizing cysteine trapping of residues at the top of transmembrane segments and within extracellular loops using cysteines in positions comprising the helix N-capping motif, and evaluating potential candidate pharmacophores directed to the natural activation pocket by attachment to a ligand that can be applied as a site-selective anchor that binds with high affinity to the receptor amino terminus. The second aim will gain insights into the receptor holo- structure by utilizing biochemical and mutagenesis strategies to define and disrupt or stabilize the interfaces between the secretin receptor amino terminus and core, and to examine functional implications. The third aim will explore the physical association between secretin and angiotensin 1a receptors, defining the structural basis and functional implications of their association, the impact of the full spectrum of types of drugs acting at each receptor, and developing strategies to take advantage of this unique hetero-receptor complex.