SPECTROSCOPY OF BIOMEMBRANE MONOLAYERS

本提案中描述的研究的总体、长期目标是 使用组合、协同、有组织的方法来研究两个-- 界面处的三维生物膜结构。具体目标 这项研究项目的特点是结合生物化学的方法， 表面化学和振动光谱学用于评价 体外生理相关研究中的结构/功能关系 肺表面活性物质模型。 这一结合的研究方法将使用生化技术来 新发现的纤溶酶原的分离与鉴定 肺表面活性物质的磷脂成分。古典朗缪尔- Blodgett表面化学技术将用于研究磷脂 表面活性物质模型络合物和隔离材料中的单层结构 来自天然的肺表面活性物质。这些实验，除了 在生理上监测磷脂的有序性和相互作用 相关的国家，将直接检验“挤出”假说 表面活性剂涂抹，即表面活性物质的一种特定成分 在连续的压缩循环中，在地表变得丰富 以产生接近于零的表面张力。红外光谱学将是 用于确定这些生物膜模型的分子构象 通过衰减全反射法在固体载体上，以及在- 通过外反射法在空气-水界面进行原位测量。我们会 监测钙离子依赖的磷脂构象，蛋白质次级 朗缪尔分子中有序多肽片段的结构和取向 红外光谱分析布洛杰特脂多肽膜。这一方法将 提供关于相互作用的分子性质的会费 磷脂、钙离子和表面活性蛋白质。我们会研究它的结构 肺表面活性物质SP-B和SP-C的空间分布 利用这些振动光谱研究双层超薄膜中的多肽 方法结合表面化学技术。我们将进一步 开发一种获取生物物理增强红外光谱的新方法 在空气-水界面处原位形成单层膜，特别是 该方法在表面活性多肽相互作用研究中的应用 具有磷脂单分子层。一种新的振动光谱方法 还将用于识别二维微域 用作超薄生物膜模型的二元混合物中的分离。这个 这些技术在体外模型系统上的结合有望 极大地提高了我们对结构和反应性的理解 活体内天然肺表面活性物质。

The general, long-term goal of the research described in this proposal is to use a combined, synergistic, approach to study organized, two- dimensional biomembrane structures at interfaces. The specific objectives of this research project are to combine the methods of biochemistry, surface chemistry, and vibrational spectroscopy for evaluation of structure/function relationships in physiologically relevant in-vitro models for pulmonary lung surfactant. This combined research approach will use biochemical techniques for the isolation and characterization of the newly-discovered plasmalogen phospholipid component of pulmonary surfactant. Classical Langmuir- Blodgett surface chemistry techniques will be used to study phospholipid monolayer structure in surfactant model complexes and in isolated material from native pulmonary surfactant. These experiments, in addition to monitoring phospholipid order and interactions in a physiologically relevant state, will directly test the "squeezing-out" hypothesis of surfactant spreading, namely that one particular component of surfactant becomes enriched at the surface during successive compression cycles, in order to produce near-zero surface tension. Infrared spectroscopy will be used to determine the molecular conformation of these biomembrane models both on solid supports via attenuated total reflectance methods, and in- situ at the air-water interface by external reflectance methods. We will monitor the Ca2+-dependent phospholipid conformation, protein secondary structure, and orientation of ordered peptide segments in Langmuir- Blodgett lipid-peptide films by infrared spectroscopy. This approach will provide dues as to the molecular nature of the interactions between phospholipids, Ca2+, and surfactant proteins. We will study the structure and spatial distribution of the pulmonary surfactant SP-B and SP-C peptides in bilayer ultrathin films using these vibrational spectroscopic methods in combination with surface chemistry techniques. We will further develop a new method for obtaining enhanced IR spectra of biophysical monolayer films in-situ at the air-water interface, and will particularly apply this method to the study of the interaction of surfactant peptides with phospholipid monolayers. A new vibrational spectroscopy approach will also be used in order to identify the two-dimensional micro-domain segregation in binary mixtures used as ultrathin biomembrane models. The combination of these techniques on in-vitro model systems promises to significantly improve our understanding of the structure and reactivity of native pulmonary surfactant in-vivo.