LIPID CONTROL OF MEMBRANE PROTEIN ORGANIZATION

这项建议涉及一个具有核心重要性的总目标， 当前的生物制药学-即确定那些结构和动态 磷脂分子的特征决定了分布， 膜结合蛋白的功能。 将努力实现这一目标 通过三个具体目标： 1)为了确定膜蛋白是否分配到 复合脂质中特定的化学结构或物理顺序 环境，并确定上述分区特性 影响膜结合酶的功能。 2)为了确定膜蛋白诱导的 磷脂分子上特定位点的微扰。 3)为了确定脂质和蛋白质中发生的构象变化， 蛋白质组分在模型囊泡系统中的相互作用 并且在相当简单的天然系统（肺表面活性剂）中， 在体外研究。 待使用的蛋白质是血型糖蛋白，其是一种充分表征的结构蛋白。 红细胞膜蛋白质和家兔CaATP酶 肌浆网，一种膜结合酶，具有几个明确的 功能协调发展的 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和差示扫描 量热法。 纯化的蛋白质将重组为二元脂质 选择混合物以模拟天然膜的相性质， 可以进行光谱分析。 平行酶活性和 物理测量将用于实现目标1。 的结构研究 用在特定位点修饰的磷脂重构的蛋白质将 用于实现目标2。 对功能至关重要的组件 肺表面活性物质将单独研究和联合使用 衰减全反射红外光谱，以了解 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).