Influence of Cholesterol on Phase Behavior and Nonrandom Mixing of Membrane Components

胆固醇对膜组分的相行为和非随机混合的影响

• 批准号: 0842839
• 负责人: Gerald Feigenson
• 金额: --
• 依托单位: Cornell University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2013-09-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0842839&HistoricalAwards=false
• 关键词: Influence; Cholesterol; Phase; Behavior; Nonrandom

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).Cholesterol is the single most abundant lipid species in mammalian plasma membranes. Nonetheless, the underlying nature of its interactions with neighboring membrane molecules has remained obscure, as have its effects on membrane biochemistry. Cholesterol is known to mix nonrandomly with other membrane lipids, and yet in most cases a clear understanding is missing for the lateral distribution of cholesterol, and the way cholesterol influences the lateral distribution of other membrane components. This project has four main conceptual elements: (i) 3-component bilayer lipid mixtures serve as models for the outer and the inner leaflets of mammalian plasma membranes. These 3-component mixtures are sufficiently complex to model real biomembranes, yet are chemically well-defined; (ii) Several independent, complementary methods are used to thoroughly map and characterize the compositional phase behavior of these model membranes; (iii) Some, perhaps all, of the most realistic model membranes have compositional regions with phase separated "nanodomains". These tiny domains, not the same as the clusters found in ordinary nonideal mixing, must be characterized as to size, and their chemical properties correlated with the well-studied macroscopic phases; (iv) Binding of membrane proteins influences the size of nanodomains, and conversely the presence of these nanodomains influences protein binding. Experimentally, this work involves large data sets of fluorescence resonance energy transfer measurements to find the lipid mixture phase boundaries and the partition behavior of fluorescent probes, including membrane-bound peptides. Confocal fluorescence microscopy is used to visualize and identify coexisting phases. X-ray diffraction experiments at the Cornell High Energy Synchrotron Source can detect nanodomains without the use of fluorescent probes. A long-range objective of this project is to discover the fundamental nature of the information that is contained in the structure of lipid polar and hydrocarbon moieties, thereby serving the broader scientific community. In this way, membrane lipids should become as well understood as are nucleic acids and amino acids. Previous work shows that lipid structural information, in part, is manifested in the phase behavior of lipid mixtures, and this project is designed to elucidate this mixture behavior. This research also will yield systematic information about properties of membrane probes that will be of wide use to the community of cell biologists who use fluorescence microscopy. As an integral feature of this project, in order to promote scientific education and training, a group of undergraduate students will receive lectures in lipid physical chemistry and fluorescence spectroscopy, followed by systematic training in lipid analytical chemistry, lipid organic synthesis, and spectroscopic and microscope techniques, which they will then use in their assigned independent research.

该奖项是根据2009年美国复苏和再投资法案（公法111-5）资助的。胆固醇是哺乳动物质膜中最丰富的单一脂质种类。 尽管如此，其与邻近膜分子相互作用的基本性质仍然不清楚，其对膜生物化学的影响也是如此。已知胆固醇与其他膜脂质非随机混合，但在大多数情况下，对胆固醇的横向分布以及胆固醇影响其他膜组分横向分布的方式缺乏清晰的理解。该项目有四个主要的概念要素：（一）3-组分双层脂质混合物作为哺乳动物质膜的外小叶和内小叶的模型。这些3-组分混合物是足够复杂的模型真实的生物膜，但化学定义良好;（ii）几个独立的，互补的方法被用来彻底映射和表征这些模型膜的组成相行为;（iii）一些，也许是所有的，最现实的模型膜的组成区域与相分离的“nanodomains”。这些微小的域，不一样的集群中发现的普通非理想的混合，必须被表征为大小，和它们的化学性质与研究的宏观相;（iv）膜蛋白的结合影响的大小的nanodomains，相反，这些nanodomains的存在影响蛋白质结合。实验上，这项工作涉及大量的荧光共振能量转移测量数据集，以找到脂质混合物相边界和荧光探针的分配行为，包括膜结合肽。共聚焦荧光显微镜用于可视化和识别共存相。康奈尔高能同步加速器源的X射线衍射实验可以在不使用荧光探针的情况下检测纳米畴。该项目的一个长期目标是发现脂质极性和碳氢化合物部分结构中所含信息的基本性质，从而为更广泛的科学界服务。这样，膜脂应该像核酸和氨基酸一样被很好地理解。以前的工作表明，脂质结构信息，在一定程度上，表现在相行为的脂质混合物，这个项目的目的是阐明这种混合物的行为。这项研究还将产生有关膜探针特性的系统信息，这些信息将广泛用于使用荧光显微镜的细胞生物学家。作为该项目的一个组成部分，为了促进科学教育和培训，一组本科生将接受脂质物理化学和荧光光谱学的讲座，然后进行脂质分析化学，脂质有机合成，光谱和显微镜技术的系统培训，然后他们将在分配的独立研究中使用。