Spectroscopy of Biomembrane Monolayers

描述（由申请人提供）：肺表面活性剂是哺乳动物肺泡的一个组成部分，用于降低表面张力和促进肺的氧摄取。缺乏表面活性剂会导致婴儿和成人的呼吸窘迫综合征。在体内，表面活性剂含有四种载脂蛋白，其中两种（SP-B和SP-C）与该材料的表面活性特性有关。SP-B已被证明可以增强吸附，而SP-C脂蛋白的α -螺旋形式已被证明可以最大限度地与脂质疏水相互作用，以增加单层的机械稳定性，并催化在空气-水界面形成3-D脂质-蛋白储层。此外，最近的研究表明，SP-C的脱酰基形式是亚稳态的，并经历了α -螺旋到β -薄片的重排，导致视觉上类似淀粉样原纤维的蛋白质聚集体。这些原纤维存在于肺泡蛋白沉积症患者的支气管肺泡液中，主要由去乙酰化的SP-C组成。分离表面活性剂的特定脂质和蛋白质成分的进展已经发展到可以利用生物物理方法研究特定结构-功能关系的地步。我们将红外光谱和光学显微镜技术应用于二维薄膜表面活性剂模型体系的研究。我们的结果已经证明了疏水表面活性剂蛋白质形成表面相关的三维颗粒的直接证据。使用一种新的计算方法，我们已经确定了SP-B和SP-C中以前未知的结构中间体，并跟踪了它们的重新定向。此外，我们已经确定pH是影响脱酰基SP-C形成淀粉样蛋白原纤维的机制之一。在下一个资助期，我们计划将这些类型的测量扩展到模拟生理相关的肺表面活性物质模型的生物物理单层。利用诱变、振动光谱学和光学显微镜的结合，我们将研究SP-B影响单层表面活性的特殊结构机制，以及脱酰基SP-C形成淀粉样原纤维的机制。

DESCRIPTION (provided by applicant): Pulmonary surfactant is an integral component of mammalian alveoli that serves to reduce surface tension and promote oxygen uptake in the lung. The lack of surfactant leads to respiratory distress syndrome in both infants and adults. In-vivo, surfactant contains four apoproteins, two of which (SP-B and SP-C) have been implicated in the surface-active properties of this material. SP-B has been shown to enhance adsorption, while the alpha-helix form of the SP-C lipoprotein has been shown to maximize hydrophobic interactions with lipids in order to increase the mechanical stability of the monolayer, and catalyze the formation of 3-D lipid-protein reservoirs at the air-water interface. Also, recent research has demonstrated that the deacylated form of SP-C is metastable and undergoes an alpha-helix to beta-sheet rearrangement resulting in protein aggregates that visually resemble amyloid fibrils. These fibrils are found in broncho-alveolar fluid isolated from patients with pulmonary alveolar proteinosis, and consist largely of deacylated SP-C. Progress in isolating specific lipid and protein components of surfactant has advanced to the point where biophysical approaches may be utilized to study specific structure-function relationships. We have applied IR spectroscopy as well as optical microscopy to the study of surfactant model systems in 2-D films. Our results have demonstrated direct evidence for the formation of surface-associated 3-D particles by hydrophobic surfactant proteins. Using a new computational approach, we have identified previously unknown structural intermediates in SP-B and SP-C and followed their reorientation. Furthermore, we have identified pH as one of the mechanisms that influence the formation of amyloid fibrils by deacylated SP-C. In the next grant period, we plan to expand these types of measurements to biophysical monolayers that mimic physiologically relevant pulmonary surfactant models. Using a combination of mutagenesis, vibrational spectroscopy and optical microscopy, we will examine the particular structural mechanisms by which SP-B influences monolayer surface activity as well as the mechanisms by which deacylated SP-C forms amyloid fibrils.