Gamma crystallin modifications & mechanisms of lens opacity

项目总结/摘要 这项建议的长期目标是确定分子机制 通过这种方式，晶体蛋白的修饰（遗传或年龄发作）导致白内障 阵这里提出的是对已知与以下疾病相关的修饰的晶体蛋白的研究： 遗传性或老年性白内障在Aim 1中，修饰的<$-晶体蛋白之间的相互作用 将单独检查（自缔合），并在目标2中修改的相互作用 将检查<$-晶体蛋白和<$-晶体蛋白。迄今为止的大多数研究都是第一次 类型，并导致更好地了解如何修改晶体蛋白自我关联 并形成导致光散射和不透明的“凝聚相”。的 第二种类型的相互作用最近出现，在维护 透镜透明度。 根据现有知识，我们提出以下假设： 晶体蛋白的修饰改变了蛋白质-蛋白质相互作用，并导致（a）自我- 突变蛋白的缔合以产生多种凝聚相，或（B）网络中的变化 突变的<$-晶体蛋白和<$-晶体蛋白之间的吸引力相互作用。目标1中的实验将 比较热力学相图，低和高分辨率结构（使用 u.v.-可见光、圆二色性、荧光、FTIR、拉曼和NMR），以及 人D-、C-和S-晶体蛋白的修饰形式与野生型蛋白的修饰形式。 相图为凝聚相的形成提供了边界条件 这导致光散射和不透明。 因此，这些研究和光散射 （静态和动态）测量是拟议研究的一个组成部分。在目标2中， 那些不自缔合的修饰形式的晶体蛋白的相互作用将是 也使用类似的技术，用天然晶状体蛋白检查。 因此，拟议的实验将确定导致 由于透镜中的晶状体蛋白修饰而导致的光散射和不透明，并且应该 提供了一个全面的了解两种形式的白内障疾病-遗传，以及 老年性白内障

Project Summary/Abstract The long-term objective of this proposal is to determine the molecular mechanisms by which modifications of the ¿-crystallins (genetic or age-onset), lead to cataract formation. Proposed here are studies on modified ¿-crystallins known to be associated with either genetic or age-onset cataract. In Aim 1 interactions between modified ¿-crystallins alone will be examined (self-association), and in Aim 2 interactions of modified ¿-crystallins with ¿-crystallin will be examined. Most studies to date have been of the first type and have led to a better understanding of how modified ¿-crystallins self-associate and form "condensed phases" which are responsible for light scattering and opacity. The second type of interaction has recently emerged as being equally important in maintaining lens transparency. Based on current knowledge, we propose the following hypothesis: Modifications of the ¿-crystallins alter the protein-protein interactions and lead to either (a) self- association of mutant proteins to yield a variety of condensed phases or (b) a change in the net attractive interactions between mutant ¿-crystallins and ¿-crystallin. Experiments in Aim 1 will compare the thermodynamic phase diagrams, the low and high-resolution structures (using u.v.-visible, circular dichroism, fluorescence, FTIR, Raman and NMR), and stabilities of the modified forms of human ¿D-, ¿C-, and ¿S- crystallins, with those of the wild-type protein. Phase diagrams provide the boundary conditions for the formation of condensed phases which lead to light scattering and opacity. Therefore, these studies and light scattering measurements (static and dynamic) are an integral part of the proposed studies. In Aim 2, the interactions of those modified forms of ¿-crystallins which do not self-associate will be examined with native ¿-crystallin, also using similar techniques. Thus, the proposed experiments will identify molecular mechanisms that lead to light scattering and opacity due to ¿-crystallin modifications in the lens, and should provide a comprehensive understanding of both forms of cataract disease - genetic, as well as age-onset cataract.