Membrane-targeting calcium sensors in vision

描述（由申请人提供）：本研究项目自成立以来的总体目标是开发核磁共振（NMR）技术，并将其与其他实验方法结合使用，以阐明参与视觉和其他信号转导过程的选定膜靶向蛋白的分子结构和生理功能。在接下来的五年里，我们将使用NMR，荧光，微量热法，X射线晶体学和高通量功能分析来描述钙-肉豆蔻酰开关蛋白家族的结构，功能和作用机制，这些蛋白在信号传导中作为膜靶向调节剂，并与视网膜和神经系统疾病有关。特别地，视网膜恢复蛋白与癌症相关的视网膜病有关，视网膜病是由另一组织中的原发性肿瘤引起的视网膜中的自身免疫性疾病。此外，鸟苷酸环化酶激活蛋白（GCAP）中的点突变与常染色体显性视锥细胞营养不良有遗传联系。通过继续深入研究视网膜恢复素和GCAP蛋白，并扩大其范围，包括同系物和目标，我们希望获得更深入的了解钙-肉豆蔻酰开关如何在膜信号传导和视网膜疾病中发挥作用。特别是，我们想了解共价连接的肉豆蔻酰基如何与钙结合位点和靶蛋白协同工作，以引导这一家族的蛋白质与膜结合的目标。具体目标是：（1）确定光激活光感受器中激活光感受器鸟苷酸环化酶的鸟苷酸环化酶的分子结构，其可用作设计预防与GCAP遗传相关的视网膜变性疾病的治疗剂的分子基础。(2)使用新开发的固态NMR技术确定与脂质双层膜结合的recoverin和靶标的结构。目的是了解膜结合引起的结构变化。（三）、阐明Ca 2 +-肉豆蔻酰开关蛋白与靶蛋白结合的分子相互作用，并发现对靶点识别和药物设计重要的原子水平结构决定因素。(4)了解钙-肉豆蔻酰开关家族的进化根源。其目的是确定钙级联G蛋白级联的耦合是否出现在进化的早期。

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.