Membrane Remodeling During Viral Infection, Parasite Inv

该项目旨在了解生理和致病事件期间膜重塑的物理化学机制。有三个组成部分： 1.大肠杆菌中毒的机制。α-溶血素是由大肠杆菌分泌的靶向真核细胞质膜的细胞外蛋白毒素（117 KDa）。我们研究了这种毒素与膜的相互作用，使用平面磷脂双层。对于所有测试的脂质混合物，除了纳摩尔浓度的毒素导致膜电导的增加和膜稳定性的降低。HlyA降低膜寿命高达三个数量级的电压？依赖的方式。使用多孔形成的理论，我们分析了这些数据，以量化HlyA如何减少膜的线张力（即形成新孔边缘所需的能量）。然而，与添加正曲率剂溶血磷脂酰胆碱将协同降低线张力的预期相反，其添加显著稳定了HlyA处理的膜。HlyA也出现在添加了它的双层膜上。这些结果导致新的考虑现有模型的蛋白质-淀粉酶孔，并导致我们提出了一个新的模型，这样的孔。 2.了解膜生物物理学。在物理学中，我们通常希望减少系统的行为，以使其符合公式。在生物学中，所有的细节都很重要：组织使它活起来。非重复聚合物生命代码动画。因此，质膜中蛋白质的组织必须是生命成功的重要组成部分。与细胞干重的三分之一是膜的事实相一致，真核基因组上大约一半的蛋白质是膜蛋白。因此，大约有一半的生物过程发生在膜上，这些过程中的每一个都有属于物理学领域的功能方面。这并不奇怪，膜生物物理学包括大约一半的主题海报和会谈会议上致力于所有生物物理学？膜包裹着几乎所有的生物。由于水是可溶性蛋白质的溶剂，磷脂双层膜是膜蛋白质的溶剂，是生物膜的基础。膜是在某些方面有序而在其他方面无序的半晶体阵列，具有流体和固体特性。它只有两个分子厚，但面积可以是平方毫米（例如，蛋）或长度为几米的圆柱体（例如，长颈鹿的轴突）。我们最接近的日常经验与薄膜是在玩肥皂泡，这是由薄膜的水内衬洗涤剂。幸运的是，磷脂双层本身对于许多脂质组合物是稳定的，并且双层在这些脂质充分水合后自组装。因此，它的性质和自我相互作用可以在没有蛋白质的情况下进行广泛的体外研究。我们对膜骨架的物理性质的理解主要来自于对双层的光谱学、显微镜和电生理学的研究。 3.蛋白质是如何形成生物形状的？生物膜呈现出各种功能相关的形状，而这些形状产生的机制在很大程度上尚不清楚。我们已经分类了可能的曲率产生机制，由脂质构成的膜双层和蛋白质相互作用，或嵌入在膜中。我们描述了膜的弹性性质，以制定蛋白质和脂质的结构和能量要求，使它们能够一起工作，产生细胞内运输过程中看到的膜形状。

This project is aimed at the understanding of the physico-chemical mechanisms of membrane remodeling during physiological and pathogenic events. There are three components: 1. The mechanism of E Coli toxicosis. alpha-Hemolysin is an extracellular protein toxin (117 KDa) secreted by Escherichia coli that targets the plasma membranes of eukaryotic cells. We studied the interaction of this toxin with membranes using planar phospholipid bilayers. For all lipid mixtures tested, addition of nanomolar concentrations of toxin resulted in an increase of membrane conductance and a decrease in membrane stability. HlyA decreased membrane lifetime up to three orders of magnitude in a voltage?dependent manner. Using a theory for lipidic pore formation, we analyzed this data in order to quantify how HlyA diminished the line tension of the membrane (i.e the energy required to form the edge of a new pore). However, in contrast to the expectation that adding the positive curvature agent lysophosphatidylcholine would synergistically lower line tension, its addition significantly stabilized HlyA-treated membranes. HlyA also appeared to thicken bilayers to which it was added. These results lead to new considerations for existing models for proteo-lipidic pores, and led to our proposing a new model for such pores. 2. Understanding membrane biophysics. In physics, we usually want to reduce the behavior of the system to fit it to a formula. In biology, all the details are important: organization makes it live. Non-repeating polymers of life code for animation. Thus the organization of proteins in the plasma membrane must be an essential part of the success of life. Consistent with fact that about a third of the dry weight of a cell is membrane, roughly half of all proteins on a eukaryotic genome are membrane proteins. Thus roughly half of biological processes occur on membranes, and each of these processes will have aspects of its function that fall into the realm of physics. It is not surprising that membrane biophysics comprises roughly half of the topics for posters and talks at meetings dedicated to all biophysics ? membranes wraps themselves around almost all of biology. As water is the solvent for soluble proteins, the phospholipid bilayer membrane is the solvent for membrane proteins and the basis of the biological membrane. A semi-crystalline array that is ordered in some aspects and disordered in other aspects, a membrane has both a fluid and a solid character. It is only two molecules thick but can be square millimeters in area (e.g., eggs) or cylinders meters in length (e.g., axons in giraffes). Our closest everyday experience with thin films comes during play with soap bubbles, which are made of thin films of water lined by detergents. Luckily, the phospholipid bilayer itself is stable for many lipid compositions, and bilayers self-assemble upon sufficient hydration of these lipids. Therefore its properties and self-interactions can be extensively studied without proteins in vitro. Our understanding of the physical nature of the membrane backbone comes mostly from studies on the spectroscopy, microscopy, and electrophysiology of bilayers. 3. How do proteins produce biological shape? Biological membranes exhibit various function-related shapes, and the mechanism by which these shapes are created is largely unclear. We have classified possible curvature-generating mechanisms that are provided by lipids that constitute the membrane bilayer and by proteins that interact with, or are embedded in, the membrane. We describe membrane elastic properties in order to formulate the structural and energetic requirements of proteins and lipids that would enable them to work together to generate the membrane shapes seen during intracellular trafficking.