Structural Dynamics of Retinal Binding and Release

视网膜结合和释放的结构动力学

• 批准号: 7435259
• 负责人: David L Farrens
• 金额: $ 25.65万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7435259
• 关键词: Affect; Apoptosis; Arrestin; Arrestins; Attenuated; Binding; Biological Assay; Centrifugation; Column Chromatography; Complex; Condition; Disease; Event; Family; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Goals; Human Genome; Hydrolysis; Kinetics; Knowledge; Label; Leber' s disease; Light; Link; Membrane; Methods; Modeling; Molecular; Monitor; Mutagenesis; Mutation; Night Blindness; Nonexudative age-related macular degeneration; Opsin; Pharmaceutical Preparations; Pharmacologic Substance; Play; Process; Proteins; Research; Resolution; Retinal; Retinitis Pigmentosa; Rhodopsin; Role; Schiff Bases; Signal Transduction; Signaling Protein; Site; Structure; System; Testing; Time; Vertebrate Photoreceptors; Vision research; Visual; Visual Signal Transduction Pathway; adduct; attenuation; chromophore; extracellular; receptor

DESCRIPTION (provided by applicant): The long-term goal of our research is to understand the molecular mechanisms through which G-protein coupled receptors (GPCRs) are activated and attenuated. These receptors represent the largest family in the human genome, and most importantly, are the target of most pharmaceutical drugs. Our studies focus primarily on the GPCR rhodopsin and its affiliate proteins. Although knowledge of the proteins involved in visual signaling has been enormously advanced through recent crystallographic studies, the critical structural changes they undergo during activation and attenuation remain largely a matter of speculation. In particular, we lack even the most rudimentary information about the dynamic events that occur during the attenuation of rhodopsin signaling, namely, the mechanisms used to release retinal from the opsin-binding pocket, and how these processes affect arrestin binding and release. Understanding these processes is of fundamental importance for vision research, as the stability of the retinal linkage varies widely among different opsins and is a factor in some visual disease states. Furthermore, little is known regarding the mechanism and kinetics of arrestin release from activated rhodopsin. In Aim I of this proposal we will determine the mechanisms through which rhodopsin controls the hydrolysis of its retinal Schiff base linkage. In Aim II we will use our expertise in site-directed labeling methods to examine dynamic and structural changes occurring in the extracellular loop region of rhodopsin as it binds and releases retinal. Finally, in Aim III, we will utilize a new assay we have developed to study arrestin release from activated rhodopsin. Understanding what makes arrestin "let go" after binding rhodopsin is fundamentally important, since stable rhodopsin-arrestin complexes have been suggested to be a contributing factor in apoptosis and autosomal dominant retinitis pigmentosa (ADRP). Furthermore, using our new assay, we find intriguing evidence that arrestin binds to and traps a post-Meta II photodecay product, possibly Meta III. Thus, arrestin may also serve to limit the release of free retinal under bright light conditions, and thus help limit the formation of oxidative retinal adducts that can contribute to diseases like atrophic age-related macular degeneration (AMD).

描述（由申请人提供）：我们研究的长期目标是了解分子机制通过激活G蛋白偶联受体（GPCR）的分子机制。这些受体是人类基因组中最大的家族，最重要的是大多数药物的靶标。我们的研究主要关注GPCR视紫红质及其关联蛋白。尽管通过最近的晶体学研究，人们对参与视觉信号的蛋白质的了解已得到了极大的提高，但它们在激活和衰减期间经历的关键结构变化在很大程度上仍然是猜测的问题。特别是，我们甚至缺乏有关在视紫红蛋白信号传导衰减期间发生的动态事件的最基本的信息，即，用于从Opsin结合口袋释放视网膜的机制，以及这些过程如何影响抑制蛋白结合和释放。理解这些过程对于视力研究至关重要，因为视网膜连接的稳定性在不同的Opsins之间差异很大，并且是某些视觉疾病状态的一个因素。此外，关于激活的视紫红质释放抑制蛋白的机制和动力学知之甚少。 在该提案的目标中，我们将确定视紫红质控制其视网膜席夫碱基的水解的机制。在AIM II中，我们将使用我们的专业知识在定位的标记方法中使用我们的动态和结构变化，因为它结合并释放了视网膜，在Rhodopsin的细胞外环区域发生。最后，在AIM III中，我们将利用我们开发的新测定方法来研究激活的视紫红质的释放。了解使抑制蛋白在结合视紫红蛋白后“放开”的原因在根本上很重要，因为稳定的视紫红质 - arrestin络合物被认为是凋亡和常染色体显性视网膜炎色素（ADRP）的促成因素。此外，使用我们的新测定，我们发现了令人着迷的证据，表明停滞蛋白与Meta II后光模型产物（可能是Meta III）结合并捕获。因此，逮捕蛋白还可以限制在明亮的光线条件下自由视网膜的释放，从而有助于限制形成氧化性视网膜加合物的形成，这可能导致诸如萎缩与年龄相关的黄斑变性（AMD）等疾病。