Endogenous G-Protein Coupled Receptor Antagonists

描述（由申请人提供）：Agouti-related protein（AGRP）和agglutamine（ASP）是迄今为止鉴定的仅有的两种天然存在的G蛋白偶联受体（GPCR）拮抗剂，当在小鼠中过度表达时，导致肥胖表型。肥胖（身体质量指数>25）困扰着美国数百万人（估计美国有9700万成年人）。是心脏病、II型糖尿病、中风和高血压的主要危险因素。在工业化国家中，肥胖问题由于暴饮暴食、高脂肪含量饮食和缺乏锻炼而变得更加复杂。在过去的几年里，已经发现了超过25种参与和调节摄食行为和能量稳态的途径。黑皮质素途径包括五个这样的遗传因素，已被证明介导体重稳态，并在修改时，导致肥胖。黑皮质素途径包括源自前激素原阿黑皮素原（POMC）基因转录物的黑皮质素激动剂、迄今为止鉴定的五种黑皮质素受体（MC 1 R-MC 5 R）以及仅有的两种天然存在的GPCR拮抗剂，即阿格列汀（ASP）和刺豚鼠相关蛋白（AGRP）。被鉴定为参与能量稳态的五种黑皮质素遗传因子是POMC、ASP、AGRP和脑特异性黑皮质素-4（MC 4 R）和黑皮质素-3（MC 3R）受体。AGRP和ASP是拮抗黑皮质素受体的旁分泌因子。具体而言，ASP拮抗皮肤黑皮质素-1受体（MC 1 R）和大脑MC 4 R。AGRP通常仅拮抗脑中的黑皮质素受体MC 3R和MC 4 R。在诊断为神经性厌食症的人类中已经鉴定出将AGRP与人类生理疾病状态联系起来的多态性。ASP和AGRP的长度均为131-132个氨基酸（取决于物种），并具有C-末端结构域，其purone含有5个二硫键。C-末端结构域已被鉴定为在黑皮质素受体上具有拮抗性质的蛋白质区域。这些数据支持AGRP和ASP参与黑皮质素受体介导的生理途径的调节的假设。本提案的研究目标是：1）鉴定对黑皮质素受体蛋白的分子识别和功能活性重要的AGRP蛋白的结构决定因素，2）鉴定推定的AGRP-MC 4受体相互作用。除了生成关于仅有的两种内源性GPCR拮抗剂之一的结构和功能信息之外，还将开发新的工具，其可以在体内使用以获得对导致肥胖和皮肤色素沉着的生理机制的更广泛的理解。

DESCRIPTION (provided by applicant): Agouti-related protein (AGRP) and agouti (ASP) are the only two naturally occurring antagonists of G protein coupled receptors (GPCRs) identified to date, and when over expressed in mice, result in an obese phenotype. Obesity (body mass index >25) afflicts millions of people in the United States (an estimated 97 million adults in the U.S.) and other countries, and is a major risk factor for heart disease, type II diabetes mellitus, stroke and hypertension. In industrialized countries, the problem of obesity is compounded by overeating, a high fat content diet, and a lack of exercise. The last few years have seen the characterization of over 25 pathways that have been identified to participate in and regulate feeding behavior and energy homeostasis. The melanocortin pathway includes five such genetic factors that have been demonstrated to mediate weight homeostasis, and when modified, result in obesity. The melanocortin pathway includes the melanocortin agonists, derived from the preprohormone proopiomelanocortin (POMC) gene transcript, the five melanocortin receptors identified to date (MC1R-MC5R), and the only 2 naturally occurring antagonists of GPCRs, agouti (ASP) and agouti-related protein (AGRP). The five melanocortin genetic factors identified as being involved in energy homeostasis are POMC, ASP, AGRP, and the brain specific melanocortin-4 (MC4R) and the melanocortin-3 (MC3R) receptors. AGRP and ASP are paracrine factors that antagonize the melanocortin receptors. Specifically, ASP antagonizes the skin melanocortin-1 receptor (MC1R) and the brain MC4R. AGRP normally only antagonizes the melanocortin receptors in the brain, MC3R and MC4R. A polymorphism has been identified in humans diagnosed with anorexia nervosa, linking AGRP with a human physiological disease state. Both ASP and AGRP are 131-132 amino acids in length (depending upon species), and possess a C-terminal domain that putatively contains 5 disulfide bridges. The C-terminal domain has been identified as the region of the protein that possesses antagonistic properties at the melanocortin receptors. These data support the hypothesis that AGRP and ASP are involved in the regulation of melanocortin receptor mediated physiological pathways. The research objectives of this proposal are to 1) identify structural determinants of the AGRP protein important for molecular recognition and functional activity at melanocortin receptor proteins, and 2) identify putative AGRP-MC4 receptor interactions. In addition to generating structural and functional information about one of the only two endogenous GPCR antagonists, new tools will be developed which may be utilized in vivo to gain a broader understanding of the physiological mechanism(s) resulting in obesity and skin pigmentation.