MEMBRANE BASIS OF VISUAL EXCITATION

视觉兴奋的膜基础

• 批准号: 6986060
• 负责人: Michael F Brown
• 金额: $ 33.11万
• 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 2008-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6986060
• 关键词: biological signal transduction; biophysics; cell surface receptors; chromophore; computer simulation; conformation; lipid bilayer membrane; low angle X ray diffraction analysis; membrane activity; membrane lipids; molecular dynamics; nuclear magnetic resonance spectroscopy; photochemistry; protein structure function; rhodopsin; visual stimulus

DESCRIPTION (provided by applicant): The scientific focus of this research is to characterize the events at the molecular level which lead to the triggering of visual signal transduction by rhodopsin, a G protein-coupled receptor. The new biophysical knowledge to be obtained is significant from a fundamental standpoint, and also with regard to visual diseases such as retinitis pigmentosa. A combined structure-function approach will elucidate the membrane basis of visual signalling. Rhodopsin is an integral membrane protein, and is part of a supramolecular assembly comprising a polyunsaturated lipid bilayer, with a characteristic polar head group composition. As a result, the structural properties of rhodopsin as well as the membrane lipid constituents are both important for understanding the mechanism of visual signal transduction. To test the above conceptual framework, we shall mainly employ a key biophysical methodology, deuterium (2H) NMR spectroscopy. First, we plan to use solid state 2H NMR technology to investigate recombinant membranes in which rhodopsin has a deuterated retinal chromophore. The local conformation, orientation, and mobility of retinal will be studied in the dark-adapted ground state of rhodopsin in conjunction with the recently published X-ray crystal structure. Novel solid-state NMR methods will include lineshape simulations of semi-random distributions of aligned membranes, together with relaxation measurements. Next, solid-state 2H NMR spectroscopy of aligned membrane samples will be used to investigate the retinal chromophore in the bathorhodopsin, Meta I, and Meta II states. An innovative new aspect is to elucidate the changes that accompany the Meta I-Meta II transition of rhodopsin, the trigger for visual signal transduction. Third, molecular dynamics computer technology will be developed and applied to investigate membrane lipid-rhodopsin interactions in conjunction with the rhodopsin 3-D structure. Additional research will use 2H and 31P NMR spectroscopy of the polyunsaturated membrane lipids to distinguish alternative hypotheses for their influences on the Meta I-Meta II conformational transition. Our hypothesis is that the transition involves a change in the curvature elastic stress/strain of the lipid bilayer, thus providing for a coupling of the protein energetics to the properties of non-lamellar forming membrane lipids. Lastly, 2H NMR spectroscopy of bilayers and non-lamellar phases of polyunsaturated membrane lipids will investigate their equilibrium and dynamical properties in relation to rhodopsin function. Thus we 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 G protein-coupled receptors and signal transduction in general.

描述（由申请人提供）：本研究的科学重点是在分子水平上表征导致视紫红质（一种G蛋白偶联受体）触发视觉信号转导的事件。从基本观点来看，获得的新的生物物理知识是重要的，对于视网膜色素变性等视觉疾病也是如此。结合结构-功能的方法将阐明视觉信号的膜基础。视紫红质是一种完整的膜蛋白，并且是包含具有特征性极性头基组成的多不饱和脂质双层的超分子组装体的一部分。因此，视紫红质的结构特性以及膜脂成分对于理解视觉信号转导机制都是重要的。为了测试上述概念框架，我们将主要采用一种关键的生物物理方法，氘（2 H）NMR光谱。首先，我们计划使用固态2 H NMR技术来研究重组膜，其中视紫红质具有氘代视网膜发色团。局部构象，取向和流动性的视网膜将研究在黑暗中适应的视紫红质结合最近公布的X射线晶体结构的基态。新的固态NMR方法将包括线形模拟对齐膜的半随机分布，以及弛豫测量。接下来，对齐的膜样品的固态2 H NMR光谱将用于研究视紫红质、Meta I和Meta II状态中的视网膜发色团。一个创新的新的方面是阐明的变化，伴随着Meta I-Meta II转换的视紫红质，触发视觉信号转导。第三，分子动力学计算机技术将被开发和应用于研究膜脂视紫红质的相互作用与视紫红质的三维结构。更多的研究将使用2 H和31 P NMR光谱的多不饱和膜脂，以区分其对Meta I-Meta II构象转变的影响的替代假设。我们的假设是，过渡涉及的脂质双层的曲率弹性应力/应变的变化，从而提供了一个耦合的蛋白质能量的非层状形成膜脂质的属性。最后，2 H NMR光谱的双层和非层状相的多不饱和膜脂质将调查其平衡和动力学性质的关系，视紫红质功能。因此，我们打算提供一个全面的图片如何视紫红质与双层脂质产生触发视觉兴奋的脊椎动物杆，这是一个范例，G蛋白偶联受体和信号转导一般。