RAMP-altered Class B GPCR Hormone Recognition

描述（由申请方提供）：人降钙素（CTR）和降钙素样（CGRP）受体是细胞表面B G类蛋白偶联受体（GPCR），与受体活性修饰蛋白（RAMP）共受体相关，介导降钙素家族肽激素的信号传导。肽肾上腺髓质素（AM）、降钙素基因相关肽（CGRP）和胰淀素（AMY）的不同生物学作用是通过它们与由RAMP或CTR和三种RAMP之一组成的特定分子复合物结合而引起的。AM和CGRP分别通过CLR-RAMP 2或3和CLR-RAMP 1复合物的信号传导来调节心血管系统。AMY通过与RAMP 1、2或3相关的CTR信号传导调节血糖水平。CTR-RAMP复合物是重要的药物靶点。靶向CLR-RAMP复合物的治疗剂具有治疗急性心肌梗死、肺动脉高压、偏头痛、癌症和几种其它病症的潜力。CTR-RAMP复合物是糖尿病药物SymlinTM的靶点。尽管靶向RAMP/CTR-RAMP复合物的临床价值，但肽激素识别和RAMP调节RAMP/CTR激素特异性的分子机制知之甚少，这在很大程度上是因为缺乏肽-受体复合物的结构信息。激素结合亲和力和特异性在很大程度上由β-内酰胺酶、CTR和RAMP整合膜蛋白的胞外结构域（ECD）决定。本提案的目标是确定AM、CGRP和AMY与其各自的EGFR/CTR ECD-RAMP ECD复合物结合的晶体结构，以定义确定选择性肽结合的精确分子结构。此外，我们将通过生物化学和基于细胞的药理学实验将结构和功能联系起来。我们提出了以下三个目标：（1）确定人CLR-RAMP 2 ECD复合物识别AM的机制。(2)确定RAMP 1如何将CGRP选择性赋予CLR-RAMP 1 ECD复合物。(3)确定人CTR-RAMP 1 ECD复合物识别AMY的机制。实现这些目标将定义RAMP改变的B类GPCR激素识别的分子基础，并提供结构模板来指导靶向受体的治疗剂的合理设计。 公共卫生相关性：该项目研究肽激素如何与人体细胞表面的受体结合。我们研究的激素是心血管系统和血糖水平的重要调节剂。这些受体是一个被称为GPCR的受体大家族的成员，该家族几乎调节人类生理学的所有方面。发现这些激素如何与其受体结合是很重要的，因为它将有助于开发针对心血管疾病和糖尿病受体的药物。这些研究对于我们将获得的关于GPCR的一般知识也很重要，这些知识将广泛适用于人类健康和疾病的几个领域。

DESCRIPTION (provided by applicant): The human calcitonin (CTR) and calcitonin-like (CLR) receptors are cell-surface class B G protein-coupled receptors (GPCRs) that associate with receptor activity modifying protein (RAMP) co-receptors to mediate signaling by calcitonin family peptide hormones. The distinct biological actions of the peptides adrenomedullin (AM), calcitonin gene-related peptide (CGRP), and amylin (AMY) are elicited by their binding to specific molecular complexes comprised of CLR or CTR and one of three RAMPs. AM and CGRP regulate the cardiovascular system by signaling through CLR-RAMP2 or 3 and CLR-RAMP1 complexes, respectively. AMY regulates blood glucose levels by signaling through CTR associated with RAMP1, 2, or 3. CLR/CTR-RAMP complexes are important drug targets. Therapeutic agents targeting CLR-RAMP complexes have the potential to treat acute myocardial infarction, pulmonary hypertension, migraine headache, cancer, and several other disorders. CTR-RAMP complexes are targets of the diabetes drug SymlinTM. Despite the clinical value of targeting CLR/CTR-RAMP complexes, the molecular mechanisms of peptide hormone recognition and RAMP modulation of CLR/CTR hormone specificity are poorly understood, in large part because of a lack of structural information for the peptide-receptor complexes. Hormone binding affinity and specificity are largely determined by the extracellular domains (ECDs) of the CLR, CTR, and RAMP integral membrane proteins. The goals of this proposal are to determine crystal structures of AM, CGRP, and AMY bound to their respective CLR/CTR ECD-RAMP ECD complexes in order to define the precise molecular architectures that determine selective peptide binding. In addition, we will correlate structure and function through biochemical and cell-based pharmacological experiments. We propose the following three aims: (1) Determine the mechanism of AM recognition by the human CLR-RAMP2 ECD complex. (2) Determine how RAMP1 confers CGRP selectivity to the CLR-RAMP1 ECD complex. (3) Determine the mechanism of AMY recognition by the human CTR-RAMP1 ECD complex. Achieving these aims will define the molecular bases for RAMP-altered class B GPCR hormone recognition and provide structural templates to guide the rational design of therapeutic agents targeting the receptors. PUBLIC HEALTH RELEVANCE: This project addresses how peptide hormones bind to receptors on the surface of human cells. The hormones we study are important regulators of the cardiovascular system and blood glucose levels. The receptors are members of a large family of receptors known as GPCRs that regulate virtually all aspects of human physiology. Discovering how these hormones bind to their receptors is important because it will aid the development of drugs that target the receptors for the treatment of cardiovascular disorders and diabetes. These studies are also important for the general knowledge we will gain about GPCRs, which will be broadly applicable to several areas of human health and disease.