High Resolution Membrane Structure From Fluorescence

该项目的目标将是继续研究脂筏的结构和功能： 在哺乳动物和其他真核细胞中发现的鞘磷脂和富含胆固醇的膜域。其中 其他功能，木筏涉及膜之间蛋白质和脂类的分类，信号转导， 以及某些类型的细菌和病毒感染。普雷沃乌斯在这个项目中的研究，作为一个(继续)的一部分 与黛博拉·布朗(Stony Brook)博士的实验室合作，建立了一些基本原则 解释鞘脂、甾醇和某些蛋白质是如何形成木筏的。新的荧光和荧光 允许检测纳米级木筏并完全控制木筏成分的淬火方法 利用以前开发的光谱和生化技术，将被用来定义几个 仍然神秘的规则控制着木筏中的脂肪和蛋白质的参与。浮筏形成的基础 贫鞘磷脂质膜内叶将在模型膜系统中进行研究。木筏成型 将研究胆固醇的生物合成前体的行为，以帮助确定疾病是如何阻止步骤的 胆固醇的生物合成(例如Smith-Lemli-Opitz病)可能与RAFT的有害变化有关 行为。神经酰胺取代了木筏上的甾醇。出于这个原因，生物上重要的富含神经酰胺的木筏 将在模型系统和单元中进行研究，以确定它们在性能方面与普通木筏的不同之处 和蛋白质的相互作用。为了获得对筏子形成原理的更多见解，各种小的 具有RAFT促进和RAFT不稳定行为的分子将在模型膜和细胞中进行研究。 作为这些研究的一部分，已知的多烯的浮筏不稳定效应的功能意义 抗生素将在模型膜和细胞中进行研究。蛋白质与木筏的相互作用将是 研究确定蛋白质结构与RAFT亲和力之间的关系。脂质锚定的跨膜 和胆固醇结合蛋白将被比较。已发现的支持筏子形成的甾醇类似物对不同 上一次授权期的学位将被用来定义蛋白质固醇结合专一性的性质。是否 蛋白质与普通和富含神经酰胺的木筏的相互作用也将被确定。最后，学位 将研究哪些蛋白质可以调节RAFT的形成。

The goal of this project will be to continue studies of the structure and function of lipid rafts: ordered sphingolipid and cholesterol-rich membrane domains found in mammalian and other eukaryotic cells. Among other functions, rafts are implicated in sorting of proteins and lipids between membranes, signal transduction, and some types of bacterial and viral infections. Prevoius studies in this project, as part of a (continuing) collaboration with the lab of Dr. Deborah Brown (Stony Brook), established some of the basic principles explaining how sphingolipids, sterols and certain proteins form rafts. New fluorescence and fluorescence quenching methods allowing detection of nanoscale rafts with full control over raft composition, combined with previously developed spectroscopic and biochemical techniques, will be used to define several of the still mysterious rules controlling lipid and protein participation in rafts. The basis of raft formation in the sphingolipid-poor plasma membrane inner leaflet will be studied in model membrane systems. Raft-forming behavior of biosynthetic precursors of cholesterol will be studied to help define how diseases blocking steps in cholesterol biosynthesis (e.g. Smith-Lemli-Opitz disease) may be related to deleterious changes in raft behavior. Ceramide displaces sterols from rafts. For this reason, biologically important ceramide-rich rafts will be studied in model systems and cells to define how they differ from ordinary rafts in terms of properties and protein interactions. To gain additional insights into the principles of raft formation, various small molecules with raft-promoting and raft-destabilizing behaviors will be studied in model membranes and cells. As part of these studies, the functional significance of the known raft-destabilizing effects of polyene antibiotics will be studied in both model membranes and cells. The interaction of proteins with rafts will be studied to define the relationship between protein structure and raft affinity. Transmembrane, lipid-anchored and cholesterol-binding proteins will be compared. Sterol analogs found to support raft formation to different degrees in the last grant period will be used to define the nature of protein sterol binding specificity. Whether proteins interact differently with ordinary and ceramide-rich rafts will also be determined. Finally, the degree to which proteins can regulate raft formation will be studied.