LIPID CONTROL OF MEMBRANE PROTEIN ORGANIZATION

这项建议涉及在#年具有核心重要性的总体目标 当前的生物物理学--即确定那些结构和动力学 控制分布和分布的磷脂分子的特征 膜结合蛋白的功能。这一目标将得到实现。 通过三个具体目标： 1)确定膜蛋白是否划分为 复合脂中特殊的化学结构或物理顺序 环境，并确定上述分区特征如何 影响膜结合酶的功能。 2)确定膜蛋白诱导的大小和性质 磷脂分子上特定位置的微扰。 3)确定脂质和蛋白质的构象变化 模型囊泡系统中蛋白质组分相互作用的研究 在相当简单的天然系统(肺表面活性物质)中 在体外研究。 将使用的蛋白质是血糖素，一种具有良好特性的结构蛋白 红细胞膜蛋白和兔CAATPase 肌浆网，一种与膜结合的酶，有几个定义明确的 功能。 用于解决特定目标的物理方法是拉曼和 傅里叶变换红外光谱与差示扫描 量热法。纯化的蛋白质将被重组为二元脂类 选定的混合物以模拟天然膜的相性质，并 要服从光谱分析。平行酶活性和 将使用物理测量来实现目标1。结构研究 在特定位置修饰磷脂重组的蛋白质将 用于解决目标2。对功能至关重要的组件 肺表面活性物质将单独和结合使用进行研究 衰减全反射红外光谱分析 结构/功能关系(目标3)。

This proposal addresses a general objective of central importance in current biophysics-namely to determine those structural and dynamic features of phospholipid molecules that govern the distribution and function of membrane-bound proteins. This objective will be approached through three specific aims: 1) To determine whether membrane proteins partition into regions of particular chemical structure or physical order in complex lipid environments, and to determine how the above partitioning characteristics affect the function of membrane-bound enzymes. 2) To determine the magnitude and nature of membrane protein-induced perturbation at specific sites on phospholipid molecules. 3) To determine the conformational changes that occur in both the lipid and protein components upon their mutual interaction in model vesicle systems and in a reasonably simple native system (pulmonary surfactant) to be studied in vitro. The proteins to be used are glycophorin, a well-characterized structural protein from the erythrocyte membrane, and CaATPase from rabbit sarcoplasmic reticulum, a membrane-bound enzyme with several well-defined functions. The physical methods to be used to address the specific aims are Raman and Fourier Transform Infrared spectroscopies and Differential Scanning Calorimetry. Purified proteins will be reconstituted into binary lipid mixtures selected to mimic the phase properties of natural membranes and to be amenable to spectroscopic analysis. Parallel enzyme activity and physical measurement will be used to address Aim 1. Structural studies of proteins reconstituted with phospholipids modified at particular sites will be used to address Aim 2. The components crucial to the function of pulmonary surfactant will be studied individually and in combination using attenuated total reflectance IR spectroscopy in order to understand structure/function relationships (Aim 3).