Lipid Control of Membrane Protein Organization

描述(申请人提供)：本实验室的长期目标是确定脂/蛋白相互作用在肺表面活性物质功能中的作用。最近的兴趣是确定各种脂质物种在皮肤通透性屏障结构中的作用。将实现两个具体目标，每个目标都有与这些目标相关的物理和生物/生物医学部分： 目的1：物理：扩展红外反射吸收光谱(IRRAS)，用于原位测定A/W界面脂单分子膜的三维结构元素。 生物学：验证疏水性表面活性蛋白SP-B和C促进多层膜的形成/稳定的假设，这些多层膜在肺压缩-扩张周期中充当表面活性物质的储存库。作为对IRRAS的补充，将使用布鲁斯特角度显微镜、压力-面积等温线和脉动气泡表面测量法来表征这些薄膜。合成多肽的同等实验将有助于确定多肽作为有效的呼吸窘迫综合征等疾病治疗剂所需的结构特征。我们还将评估肺表面活性物质SP-A类似物与脂质和其他配体的相互作用，以确定其在肾小管髓鞘形成和宿主防御中的作用。 目的2：物理：应用3种新的振动显微成像技术(红外显微成像、共聚焦拉曼显微镜和衰减全反射成像)获得1.5 mm到4 mm距离尺度的空间分辨分子结构信息。这些将被用来研究皮肤屏障模型系统中类脂结构域的形成。本实验旨在了解不同种类神经酰胺在屏障功能中的作用。 生物学：利用这些方法研究皮肤分子组成的三维空间分布以及渗透促进剂和药物载体的渗透途径。最后，振动光谱对分子结构的敏感性将被用来描绘渗透促进剂对脂质构象和蛋白质二级结构的影响。

DESCRIPTION (provided by applicant): A long-term goal of this laboratory has been to determine the role of lipid/protein interaction in the function of pulmonary surfactant. A more recent interest has been to determine the roles of the various lipid species in the structure of the barrier to permeability in skin. Two specific aims, each with a physical and biological/biomedical component related to these goals, will be pursued: Aim 1: Physical: To extend infrared reflection-absorption spectroscopy (IRRAS), for determination of elements of the 3-dimensional structure of lipid monolayers in situ at the A/W interface. Biological: To test the hypothesis that the hydrophobic surfactant proteins SP-B and C facilitate the formation/stabilization of multilayers that act as reservoirs for surface-active material during lung compression-expansion cycles. To complement IRRAS, Brewster angle microscopy, pressure-area isotherms, and pulsating bubble surfactometry will be used to characterize these films. Equivalent experiments with synthetic peptides will help define structural characteristics needed for peptides to be useful therapeutic agents for diseases such as respiratory distress syndrome. We will also evaluate the interactions of lung surfactant SP-A analogues with lipids and other ligands to determine its role in tubular myelin formation and in host defense. Aim 2: Physical: To apply 3 novel vibrational microscopic imaging technologies (IR microscopic imaging, confocal Raman microscopy, and attenuated total reflectance imaging) to acquire spatially resolved molecular structure information at distance scales from 1.5 mm to 4 mm. These will be used to study lipid domain formation in model systems for the skin barrier. The experiment aims to understand the role of the various ceramide species in barrier function. Biological: To use these methods to study the 3-D spatial distribution of the molecular constituents of skin as well as the permeation pathways of penetration enhancers and drug delivery vehicles. Finally, the sensitivity of vibrational spectra to molecular structure will be used to map the effects of penetration enhancers on lipid conformation and protein secondary structure.