CATARACT RELATED MODIFICATIONS OF LENS CRYSTALLINS

描述：拟议的调查将使用新的分析方法 基于质谱学鉴定人类晶状体蛋白质和 描述他们的翻译后修饰。在第一层 分析中，两种新型的质谱学，电喷雾电离 (ESIMS)和基质辅助激光解吸(MALDI)，将用于 准确测定完整蛋白质的相对分子质量 0.01%。在第二层次的分析中，晶状体蛋白将被消化 这些多肽的分子量将直接由 联用高效液相色谱质谱 (高效液相色谱/质谱仪)。年轻、清晰的镜片结果将被用来制作数据 由于晶状体老化和白内障而导致的晶状体倾斜 比较一下。目前，来自Young的α-晶体蛋白的参考数据 旧的透明镜片已经接近完成，大量的数据已经 收集的是贝塔和伽马晶体蛋白。《公约》的具体目标 目前的建议是1)。完成贝塔和贝塔的鉴定 在人类晶状体中表达并定义后- 这些蛋白质的翻译修饰与衰老有关。2)。 确定晶状体蛋白的成分和翻译后修饰 不溶于水，可溶于盐酸胍的部分 清除来自年轻和老年捐赠者的镜片。3)。确定翻译后 与α-、β-和γ-晶体蛋白相关的修饰 白内障的早期阶段。4)。通过协作研究，确定 晶体蛋白组分与个体的翻译后修饰 用双向凝胶电泳法从人晶状体中分离出斑点。

DESCRIPTION: The proposed investigation will use new analytical methods based on mass spectrometry to identify human lens proteins and characterize their post-translational modifications. In the first level of analysis, two new types of mass spectrometry, electrospray ionization (ESIMS) and matrix-assisted laser desorption (MALDI), will be used to determine the molecular weights of intact proteins with an accuracy to 0.01%. In the second level of analysis, crystallins will be digested into peptides whose molecular weights will be determined by directly coupled high performance liquid chromatography mass spectrometry (HPLC/MS). Results for young, clear lenses will be used to make a data bank, to which results from aged and cataractous lenses will be compared. At present, reference data for alpha-crystallins from young and old clear lenses are nearly complete and considerable data have been collected for beta and gamma-crystallins. The Specific Aims of the present proposal are 1). To complete the identification of beta- and gamma-crystallins that are expressed in the human lens and define post- translational modifications of these proteins associated with aging. 2). Identify the crystallin components and post-translational modifications of the water-insoluble, guanidine hydrochloride-soluble fraction of clear lenses from young and old donors. 3). Identify post-translational modifications of alpha-, beta-, and gamma-crystallins associated with early stages of cataract. 4). Through collaborative studies, identify the crystallin components and post-translational modifications of individual spots isolated from human lenses by 2-D gel electrophoresis.