Membrane-targeting calcium sensors in vision

描述(申请人提供)：这项研究计划自开始以来的总体目标是开发核磁共振技术，并将其与其他实验方法结合使用，以阐明涉及视觉和其他信号转导过程的选定膜靶向蛋白质的分子结构和生理功能。在接下来的五年里，我们将使用核磁共振、荧光、微量热、X射线结晶学和高通量功能分析来描绘钙-肉豆蔻基开关蛋白家族的结构、功能和作用机制，这些蛋白在信号转导中作为膜靶向调节因子，与视网膜和神经疾病有关。特别是，视网膜恢复素被认为与癌症相关的视网膜病变有关，这是一种由另一组织中的原发肿瘤引起的视网膜自身免疫性疾病。此外，鸟苷酸环化激活蛋白(GCAPs)的点突变与常染色体显性遗传性视锥细胞营养不良有关。通过继续我们对视网膜恢复素和GCAP蛋白的深入研究，并将其范围扩大到包括同系物和靶点，我们希望对钙-肉豆蔻基开关如何在膜信号和视网膜疾病中发挥作用有更深入的了解。特别是，我们想了解共价连接的肉豆蔻基是如何与钙结合位点和靶蛋白协同工作的，以引导这一蛋白家族与膜结合的靶蛋白结合。其具体目的是：(1)确定光感受器中激活光感受器鸟苷环化酶的分子结构，为设计预防与GCAPs基因相关的视网膜退行性疾病的治疗药物提供分子基础。(2)利用最新发展的固体核磁共振技术确定与脂质双层膜结合的回收率和靶标的结构。目的是了解膜结合引起的结构变化。(3)。阐明与靶蛋白结合的钙-肉豆蔻酰基开关蛋白的分子相互作用，并发现对靶识别和药物设计重要的原子水平结构决定因素。(4)了解钙-肉豆蔻酰基开关家族的进化根源。其目的是确定钙级联与G蛋白级联是否在进化早期就出现了。

DESCRIPTION (provided by applicant): The overall objectives of this research program since its inception have been to develop nuclear magnetic resonance (NMR) techniques and use them in concert with other experimental approaches to elucidate the molecular structure and physiologic functions of selected membrane-targeting proteins involved in vision and other signal transduction processes. During the next five years, we will use NMR, fluorescence, microcalorimetry, x-ray crystallography and high-throughput functional analysis to delineate the structure, functions and mechanism of action of a family of calcium-myristoyl switch proteins that serve as membrane targeting regulators in signaling and are linked to retinal and neurological diseases. In particular, retinal recoverin has been implicated in cancer associated retinopathy, an autoimmune disease in the retina caused by a primary tumor in another tissue. Also, point mutations in the guanylate cycalse activating proteins (GCAPs) are genetically linked to autosomal dominant cone dystrophy. By continuing our intensive study of retinal recoverin and the GCAP proteins and by broadening its scope to encompass homologs and targets, we hope to gain a deeper understanding of how calcium-myristoyl switches operate in membrane signaling and retinal disease. In particular, we want to understand how covalently attached myristoyl groups work in concert with calcium-binding sites and target proteins to guide this family of proteins to membrane-bound targets. The specific aims are: (1) Determine the molecular structure of guanylate cyclase activating photoreceptor guanylate cyclases in light-activated photoreceptors, which may serve as a molecular basis for designing therapeutics that prevent retinal degenerative diseases genetically linked to GCAPs. (2) Determine the structure of recoverin and targets bound to lipid bilayer membranes using newly developed solid-state NMR technology. The aim is to understand structural changes induced by membrane binding. (3). Elucidate the molecular interactions of Ca2+-myristoyl switch proteins bound to target proteins and discover atomic-level structural determinants important for target recognition and drug design. (4) Understand the evolutionary roots of the calcium-myristoyl switch family. The aim is to determine whether the coupling of calcium cascades to G protein cascades arose early in evolution.