LIPID CONTROL OF MEMBRANE PROTEIN ORGANIZATION

这项工作的长期目标是确定脂肪和蛋白质如何 结构根据它们在层次结构中的相互交互而改变 建模日益复杂的膜系统，以及这些变化是如何 都与功能有关。新的FT-IR实验产生了定量的 关于磷脂小泡膜脂构象的信息 单层膜。所开发的协议适用于更多的 技术上困难但生理上相关的实验范式 比如活细胞。两个具体的目标，每个目标都有物理和 生物成分，计划：(目标1，物理)：确定 中1-、2-和3-键酰链构象态数 结构上无序的、与生物相关的阶段，如Lpha 或H/II。(目标1，生物)：检测定量IR构象 一种微生物(A.laidlawii)活细胞膜上的标记带 b)。完整细胞中的链构象将在不同的条件下进行研究 感兴趣的条件，包括其对改变的生长阶段的反应 和温度，以及添加功能干扰剂。(目标2， Phys.)：建立FT-IR用于分子表征 脂类和蛋白质在原位的构象和取向顺序 在A/W接口的单层中。我们已经建立了一个新颖的外部 反射(IRRAS)设备允许获取第一个 蛋白质单层红外光谱。(目标2，传记)：评估 生理性本质肺的结构/功能关系 表面活性剂体系。IRRAS提供了一种独特的“排挤”测试 肺表面活性物质功能假说。这一假设需要 表面活性剂的主要脂质成分在A/W时变得富集化 在连续的膨胀-压缩循环期间的界面(呼气- 体内的吸入循环)，以产生必要的低表面张力。 我们将检查挤出的发生，它对脂质的依赖 结构和构象，以及表面活性蛋白质的能力， 更改挤出参数。这些实验将允许理性 婴幼儿呼吸窘迫综合征治疗药物的设计 (RDS)和其成体对应的(ARDS)。

The long-term objective of this work is to determine how lipid and protein structures are altered upon their mutual interaction in a hierarchy of model membrane systems of increasing complexity, and how these alterations are related to function. Novel FT-IR experiments yield quantitative information about membrane lipid conformations in phospholipid vesicles or monolayer films. The developed protocols are applicable to more technically difficult but physiologically relevant experimental paradigms such as living cells. Two specific aims, each with physical and biological components, are planned: (AIM 1, PHYS.): To determine the number of 1-, 2-, and 3-bond acyl chain conformational states in structurally disordered, biologically relevant phases such as the Lalpha or the H/II. (AIM 1, BIO.): To detect quantitative IR conformational marker bands in the live cell membranes of a microorganism (A. laidlawii B). Chain conformations in intact cells will be studied under a variety of conditions of interest including its response to altered growth phases and temperatures, and addition of functionally disruptive agents. (AIM 2, PHYS.): To develop FT-IR for the molecular characterization of conformational and orientational order of both lipids and proteins in situ in monolayers at the A/W interface. We have built a novel external reflection (IRRAS) apparatus that has permitted acquisition of the first protein monolayer IR spectra. (AIM 2, BIO.): To evaluate structure/function relationships in the physiologically essential lung surfactant system. IRRAS provides a unique test of the "squeeze-out" hypothesis of pulmonary surfactant function. This hypothesis requires that the major lipid component of surfactant becomes enriched at the A/W interface during successive expansion-compression cycles (exhalation- inhalation cycles in vivo) to produce the requisite low surface tension. We will examine the occurrence of squeeze-out, its dependence on lipid structure and conformation, and the ability of surfactant proteins, to alter squeeze-out parameters. These experiments will permit rational design of therapeutic agents for infant respiratory distress syndrome (RDS) and its adult counterpart (ARDS).