ASYMMETRIC DISTRIBUTION OF CHOLESTEROL IN MEMBRANES

拟议中的研究将检查调节的基本因素 胆固醇在细胞内的分布。 为此，域 膜内胆固醇的结构和胆固醇的转移 将检查膜之间的间隙。 具体而言，工作将侧重于 甾醇载体蛋白SCP和SCP-2在调节 膜胆固醇结构域结构，并赋予特异性， 膜间胆固醇转移 初步证据显示， 这些蛋白质在增强甾醇转移的能力上有显著差异 显然是通过完全不同的机制。 SCP对膜甾醇结构域和甾醇交换的影响 将使用分离的SCP在体外检查生物膜之间的 和从大肠杆菌获得的重组SCP，表达 克隆SCP基因。 膜甾醇结构域和甾醇交换将是 用荧光胆固醇类似物测量，一种不需要 供体和受体膜分离，单色He/Cd激光器 激励和计算机化的多频相位和调制 荧光测定法 具体目标是审查： 1)SCP在调节胆固醇结构域中的作用 膜。 2)自发和SCP介导的甾醇转移的特异性。 3)机制（载体、碰撞转移、融合或直接作用） 膜），由此SCP可以增强和/或赋予对甾醇的特异性 运动 了解SCP在细胞内固醇运动中的作用需要一个 了解膜中胆固醇的结构域。 此域 结构可能对胆固醇转运和胆固醇代谢有重要影响。 调节微粒体酶如酰基辅酶A胆固醇酰基 转移酶 这项研究的长期目标是研究 SCP在胆固醇细胞内运动中的作用及其机制 参与胆固醇积累的过程。 了解这些 这些因素应该有助于增加我们对一些 涉及细胞中异常固醇运输和/或积累的疾病 （spur-cell anemia，neoplasia，atherosclerosis，and aging）.

The proposed research will examine fundamental factors that regulate cholesterol distribution within the cell. To this end both the domain structure of cholesterol within membranes and the transfer of cholesterol between membranes will be examined. specifically the work will focus on the role of the sterol carrier proteins, SCP and SCP-2, in regulation membrane cholesterol domain structure and in conferring specificity to intermembrane cholesterol transfer. Preliminary evidence indicates that these proteins differ markedly in their ability to enhance sterol transfer between membranes and apparently do so by entirely different mechanisms. The effect of SCPs on membrane sterol domains and on exchange of sterols between biological membranes will be examined in vitro using SCPs isolated from rat liver and recombinant SCPs obtained from E.coli, expressing the cloned SCP genes. Membrane sterol domains and sterol exchange will be measured with a fluorescent cholesterol analogue, a new assay not requiring separation of donor and acceptor membranes, monochromatic He/Cd laser excitation, and computerized multifrequency phase and modulation fluorometry. The specific aims are to examine: 1) Role of SCPs in regulating the domain structure of cholesterol within membranes. 2) Specificity of spontaneous and SCP-mediated sterol transfer. 3) Mechanism(s) (carrier, collisional transfer, fusion, or direct effect on membranes) whereby SCPs may enhance and/or confer specificity to sterol movement. Understanding the role of SCPs in intracellular sterol movement requires a knowledge of the domain structure of cholesterol in membranes. This domain structure may have important consequences for cholesterol transport and the regulation of microsomal enzymes such as acyl CoA cholesteryl acyl transferase. The long term goal of this research is to examine the role of SCPs in the intracellular movement of cholesterol and how they may be involved in the process of cholesterol accumulation. Knowledge of these factors should be helpful in increasing our understanding of a number of diseases involving abnormal sterol trafficking and/or accumulation in cells (spur-cell anemia, neoplasia, atherosclerosis, and aging).