MEMBRANE BASIS OF VISUAL EXCITATION

视觉兴奋的膜基础

• 批准号: 6350873
• 负责人: Michael F Brown
• 金额: $ 20.6万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-02-01 至 2003-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6350873
• 关键词: biological signal transduction; chromophore; conformation; fiber optics; flash photolysis; intermolecular interaction; lipid bilayer membrane; membrane lipids; membrane structure; nuclear magnetic resonance spectroscopy; optic nerve; protein structure function; receptor expression; rhodopsin; rod cell; structural biology; surface plasmon resonance; transducin; visual photoreceptor; visual phototransduction

DESCRIPTION: The major goal of this research is to investigate the molecular basis for the triggering of a visual nerve impulse by the vertebrate rod. Knowledge of these molecular mechanisms can provide a conceptual basis for eventual therapeutic intervention with regard to visual disorders. The rod comprises a supramolecular assembly, in which the retinal disk membranes contain rhodopsin, an integral membrane protein, together with characteristic bilayer lipids whose composition is tightly regulated. The key signal transducing event in vision is the metarhodopsin I to metarhodopsin II conformational transition. They will test the hypothesis that upon photoexcitation of rhodopsin, the retinal Schiff base of Lys-296 is deprotonated, leading to breakage of the salt bridge to Glu-113 , which is propagated to the cytoplasmic loops of the receptor. As a result, the Meta I-Meta II transition is sensitive to the deformation energy of the membrane lipid bilayer. Diseases such as retinitis pigmentosa (RP) may be associated with perturbation of the Meta I-Meta II equilibrium, due either to mutation of rhodopsin, or alteration of the bilayer lipid environment. Two complementary biophysical methods will be employed: deuterium (2H) NMR spectroscopy and surface plasmon resonance (SPR) spectroscopy. (i) 2H NMR spectroscopy will investigate the role of the bilayer deformation energy in enabling the Meta I-Meta II conformation change of rhodopsin to occur, due to area or curvature frustration of the membrane free energy. (ii) The retinal chromophore of rhodopsin will be 2H-labeled, and 2H NMR studies will be conducted to elucidate the conformation and orientation of the retinal chromophore in the dark state. They will then investigate the changes that occur upon photoexcitation, leading to movement of the transmembrane helices and formation of the activated Meta II state. (iii) SPR spectroscopy of rhodopsin in supported planar bilayers will be used to determine how the rigid body movement of the helices influences the cytoplasmic loops of the receptor, yielding the exposure of recognition sites for the G protein (transducin). Additional SPR studies will investigate the binding constants and signaling states of the proteins involved in amplification and quenching of the visual response. Thus they intend to provide a comprehensive picture of how rhodopsin together with the bilayer lipids yields triggering of visual excitation in the vertebrate rod, which is a paradigm for membrane structure-function relationships and signal transduction in general.

描述：本研究的主要目的是调查 视觉神经冲动触发的分子基础 脊椎动物杆。 这些分子机制的知识可以提供 视觉方面最终治疗干预的概念基础 紊乱 所述杆包括超分子组装体，其中所述超分子组装体包括： 视网膜盘膜含有视紫红质，一种完整的膜蛋白， 与其组成紧密结合的特征性双层脂质一起 监管. 视觉中的关键信号转导事件是变视紫红质 I到变视紫红质II的构象转变。 他们将测试 假设视紫红质光激发后，视网膜席夫碱 的Lys-296被去质子化，导致盐桥断裂， Glu-113，其传播到受体的细胞质环。 作为 结果，Meta I-Meta II转变对形变敏感 膜脂双层的能量。 视网膜色素变性等疾病 (RP)可能与Meta I-Meta II平衡的扰动有关， 由于视紫红质的突变或双层脂质的改变， 环境 将采用两种互补的生物物理方法： 氘（2 H）NMR光谱和表面等离子体共振（SPR） 谱 (i)2 H NMR光谱学将研究化合物的作用。 使Meta I-Meta II构象成为可能的双层变形能 视紫红质的变化发生，由于面积或曲率挫折的 膜自由能 (ii)视紫红质的视网膜发色团将是 将进行2 H标记和2 H NMR研究以阐明 在黑暗状态下，视网膜发色团的构象和取向。 然后他们将研究光激发时发生的变化， 导致跨膜螺旋的移动和跨膜螺旋的形成。 激活的Meta II状态。 (iii)固载光催化剂中视紫红质的SPR光谱研究 平面双层将被用来确定如何刚体运动的 螺旋影响受体的细胞质环，产生 G蛋白（转导素）识别位点的暴露。 额外 SPR研究将调查的结合常数和信号状态， 这些蛋白质参与视觉反应的放大和淬灭。 因此，他们打算提供一个全面的图片， 与双分子层脂质一起产生视觉刺激的触发， 脊椎动物的杆，这是膜结构功能的范例 一般的关系和信号转导。