CHARACTERIZATION OF MEMBRANE LIPID DOMAINS

在模型和生物膜中，脂质具有 存在于与其他横向域不同的横向域中， 组合中的组合。 然而， 生物膜内的磷脂分子种类不是 很好理解。 在拟议的工作中，化学成分 合成脂质混合物中的脂质相和脂质结构域， 分离的大肠杆菌脂质和完整的细胞质 E.大肠杆菌进行研究。 荧光 将使用帕里芳酰磷脂的极化来评估 存在脂相，而最近的化学交联 相邻脂质将用于确定 膜流体相脂质。 乙醇胺 物质将被胺特异性试剂交联， 二甲基辛二酰亚胺，并通过高压分析产物 液相和气液色谱法。 脂质种类，映射到 相和结构域在完整的膜，将被定位到 使用非特异性脂质转移的内或外单层 牛肝蛋白和化学标记 三硝基苯磺酸 这些研究将审查 E.脂肪酸代谢突变的大肠杆菌。 与各种脂肪酸补充剂一起生长， 这些细胞由于其脂质而产生功能缺陷， 组合物的 这里描述的化学和物理研究 将被整合到一个更清晰的膜模型中， 从长远来看，这将提供一个基础， 膜脂质拓扑结构与膜功能的关系。

In both model and biological membranes, lipids have the capability of existing in lateral domains which differ from other lateral groupings in composition. However, the arrangement of phospholipid molecular species within biological membranes is not well understood. In the proposed work, the chemical composition of lipid phases and lipid domains in synthetic lipid mixtures, isolated Escherichia coli lipids, and the intact cytoplasmic membrane of E. coli will be investigated. Fluorescence polarization of parinaroyl phospholipids will be used to assess the lipid phases present, while chemical crosslinking of nearest neighbor lipids will be used to determine the lipid arrangement of the membrane fluid phase lipids. Phosphatidylethanolamine species will be crosslinked by the amine-specific reagent, dimethylsuberimidate, and the products analyzed by high pressure liquid and gas-liquid chromatography. Lipid species, mapped to a phase and domain in intact membranes, will be localized to the inner or outer monolayer using the nonspecific lipid transfer protein from bovine liver and by chemical labelling with trinitrobenzenesulfonic acid. These studies will examine the membranes of E. coli with mutations in fatty acid metabolism. Grown with various fatty acid supplements, the membranes of these cells develop functional defects due to their lipid compositions. The chemical and physical studies described here will be integrated into a more clearly defined model of membrane structure, which, in the long term, will provide a basis to relate membrane lipid topology to membrane function.