LIPID CONTROL OF MEMBRANE PROTEIN ORGANIZATION

这项工作的长期目标是确定 蛋白质和磷脂的构象变化 他们之间的相互作用。所涉及的原则将 为理解分子的结构提供了基础 生物膜及其组织如何改变 在病理条件下。 为实现这一目标，将提出三个具体目标： (1)确定完整的膜蛋白是否 优先划分为具有特定化学结构的区域 或复杂磷脂环境中的物理顺序，以及 确定任何观察到的分区与 膜结合酶的功能。 (2)确定相互作用的地点和大小 由膜蛋白和膜活性多肽产生 磷脂分子。 (3)确定构象和取向顺序的变化 它们同时存在于磷脂和蛋白质成分中 它们在模型囊泡和平面双层膜中的相互作用 系统，并在相当简单的天然制剂(肺 表面活性剂)进行体外研究。 非物质文化遗产研究的主要物理方法 脂肪构型和蛋白质的微扰方式 二级结构为傅里叶变换红外(FT-IR) 光谱学。脂-蛋白质相互作用的热力学为 用高灵敏度差示扫描量热法进行评价 (DSC)。 要实现目标1，从兔肌浆网中提取CaATPase 被分离、提纯并重组为磷脂 被选择来模仿体内的环境。可混性和 系统的分区特性通过以下方式确定 FT-IR和DSC。目标2是通过重组 所需的含磷脂的膜蛋白或多肽 在酰基链中的特定位置上的氢化。摄动 在后一种情况下，会在特定的地点进行监测和衰减 全反射光谱。目标3由 蛋白质的类型和几何取向的测定 构象敏感型酰胺形成的多肽二级结构 I或II光谱区。 通过以上推论得出的组织原则 实验将扩展到一种简单的天然组织，肺 表面活性物质，有待体外研究。基因的分子基础 表面活性剂在空气-水界面的稳定性和铺展 将会被调查。

The long term objective of this work is to determine the conformational changes that occur in proteins and phospholipids upon their mutual interaction. The principles involved will provide a basis for understanding the molecular organization of biological membranes and how the organization may be altered during pathological conditions. Three specific aims will be addressed to achieve this objective: (1) To determine whether integral membrane proteins preferentially partition into regions of specific chemical structure or physical order in complex phospholipid environments, and to determine the relationship of any observed partitioning to the function of membrane-bound enzymes. (2) To determine the site and magnitude of the interactions produced by membrane proteins and membrane-active peptides on phospholipid molecules. (3) To determine changes in conformation and orientational order that occur in both the phospholipid and protein components upon their mutual interaction in model vesicle and planar bilayer systems, and in a reasonably simple native preparation (lung surfactant) to be studied in vitro. The main physical method to be employed to study in non- perturbative fashion both lipid configuration and protein secondary structure is Fourier Transform Infrared (FT-IR) spectroscopy. The thermodynamics of lipid-protein interactions is evaluated with high sensitivity Differential Scanning Calorimetry (DSC). To achieve Aim 1, CaATPase from rabbit sarcoplasmic reticulum is isolated, purified, and reconstituted into phospholipid environments selected to mimic those in vivo. The miscibility and partitioning characteristics of the system are determined with FT-IR and DSC. Aim 2 is achieved by reconstitution of the desired membrane protein or peptide with phospholipids deuterated at specific positions in the acyl chains. Perturbations at the specific site in the latter will be monitored with attenuated total reflectance spectroscopy. Aim 3 is addressed by determination of the type and geometric orientation of protein or peptide secondary structure via the conformation-sensitive Amide I or II spectral regions. The organizational principles deduced through the above experiments will be extended to a simple native tissue, lung surfactant, to be studied in vitro. The molecular basis of the stability and spreading of surfactant at the air-water interface will be probed.