Structural Basis of Rhodopsin Function

视紫质功能的结构基础

• 批准号: 6940666
• 负责人: JAVIER V NAVARRO
• 金额: $ 26.43万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-15 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6940666
• 关键词: X ray crystallography; biological signal transduction; enzyme linked immunosorbent assay; infrared spectrometry; microspectrophotometry; posttranslational modifications; protein purification; protein structure function; receptor coupling; rhodopsin; transducin; visual photoreceptor; visual phototransduction; visual pigments; western blottings

DESCRIPTION (provided by applicant): Rhodopsin is the photoreceptor responsible for dim light vision, and represents the paradigm for the large superfamily of G protein-coupled receptors (GPCRs). The importance of rhodopsin in vision and in medicine is demonstrated by the fact that its mutants are linked to various retinal dystrophies, and mutations causing constitutive activity of rhodopsins are linked to congenital stationary night blindness. Rhodopsin has the canonical seven transmembrane helices, which delineate the pocket for the 11-cis retinal chromophore that is bound to Lys296 via a protonated Schiff base. The X-ray structure of ground-state bovine rhodopsin has been elucidated to 2.8A resolution. Determining this high-resolution X-ray structure was a major step toward developing an understanding of rhodopsin's function; however, elucidating its mechanism of activation requires knowing the high-resolution structures of intermediates along the reaction pathway, as well as their modes of interaction with transducin. Specifically, we will ask how does retinal photoisomerization trigger protein structural changes that lead to activation of transducin, thereby initiating the sensation of vision. Our major hypothesis is that light triggers the rapid isomerization of the retinal, a change which the protein both accommodates and subsequently amplifies into the larger structural changes required for signaling. We are in a unique position to address this hypothesis, as we have recently achieved a higher resolution structure of rhodopsin and have for the first time identified internal water molecules unambiguously. Furthermore, we have trapped photo-intermediates of rhodopsins in the crystals and shown that they diffract. The aims of this project are to map, at unprecedented spatial and temporal resolution, the structural transformations that occur upon activation of rhodospin by solving the X-ray structures of the photo-intermediates bathorhodopsin, lumirhodopsin and metarhodopsin (The latter is the signaling state of rhodopsin). Most importantly, we will elucidate the crystal structure of the metarhodopsin-transducin complex in order to identify the interactions at the interface of this signaling unit. Accomplishing these Specific Aims will define the physical and chemical basis for the activation of rhodopsin. Furthermore, structural studies at high resolution will open the way for understanding the molecular basis for the unique photobleaching properties of rod and cone visual pigments.

描述（由申请人提供）：视紫红质是负责微光视觉的光感受器，代表G蛋白偶联受体（GPCR）大超家族的范例。视紫红质在视觉和医学中的重要性通过其突变体与各种视网膜营养不良有关的事实证明，并且引起视紫红质组成性活性的突变与先天性静止性夜盲症有关。视紫红质具有典型的七个跨膜螺旋，其描绘了通过质子化席夫碱与Lys 296结合的11-顺式视网膜发色团的口袋。基态牛视紫红质的X射线结构已被解析到2.8A的分辨率。确定这种高分辨率的X射线结构是理解视紫红质功能的重要一步;然而，阐明其激活机制需要知道反应途径沿着中间体的高分辨率结构，以及它们与转导蛋白的相互作用模式。具体来说，我们将问视网膜光异构化如何触发蛋白质结构的变化，导致激活transducin，从而启动视觉的感觉。我们的主要假设是，光触发了视网膜的快速异构化，蛋白质既适应这种变化，又随后放大成信号传导所需的更大的结构变化。我们处于一个独特的位置来解决这一假设，因为我们最近已经实现了更高分辨率的视紫红质结构，并首次明确确定了内部水分子。此外，我们已经捕获了晶体中的视紫红质的光中间体，并表明它们是可逆的。该项目的目的是通过解决光中间体bathorhodopsin，lumirhodopsin和metarhodopsin（后者是视紫红质的信号状态）的X射线结构，以前所未有的空间和时间分辨率映射rhodospin激活后发生的结构转变。最重要的是，我们将阐明的晶体结构的后视紫红质transducin复合物，以确定在这个信号单元的接口的相互作用。实现这些特定目标将定义视紫红质激活的物理和化学基础。此外，高分辨率的结构研究将为了解视杆细胞和视锥细胞独特光漂白特性的分子基础开辟道路。