MITOCHONDRIAL PROTEOMICS OF AGING

描述（由申请人提供）：本提案的目的是全面检查衰老对小鼠线粒体蛋白质组的影响。我们选择集中在线粒体蛋白质组有两个原因。首先，几乎一致同意线粒体很可能在衰老中发挥重要作用，无论是通过功能的变化（例如，减少能量产生或增加细胞凋亡）或通过ROS的产生，ROS又损伤分子导致衰老。其次，线粒体蛋白质组有一个可管理的蛋白质数量（1000至2000），使用目前的蛋白质组学技术进行研究。 虽然有大量的数据，线粒体功能随着年龄的增长而恶化，但很少有关于这种现象的分子基础的信息。我们推测，线粒体蛋白质组的改变有很高的概率在线粒体功能的年龄相关变化中发挥重要作用。我们建议通过研究从小鼠的各种组织中分离的线粒体，并确定饮食限制（DR）的影响，延缓衰老，对线粒体蛋白质组中观察到的年龄相关变化进行首次详细研究，以验证这一假设。本项目的具体目的是分析年龄和DR对1）线粒体蛋白质组中蛋白质表达，2）线粒体磷酸化蛋白质组的磷酸化，3）线粒体蛋白质的氧化修饰和4）线粒体功能的影响。这些研究将使用我们专门开发的一系列新的蛋白质组学方法进行，用于检查蛋白质表达，酶促翻译后修饰和蛋白质氧化。认识到不同的蛋白质经常看到多维色谱和凝胶电泳，我们设计的方法，使我们能够采用这两种技术在我们的研究线粒体蛋白质组。

DESCRIPTION (provided by applicant): The goal of this proposal is to conduct a comprehensive examination of the effect of aging on the mitochondrial proteome of mice. We have chosen to focus on the mitochondrial proteome for two reasons. First, there is almost unanimous agreement that mitochondria are very likely to play an important role in aging, either through changes in function (e.g., reduced energy generation or increase apoptosis) or through the generation of ROS, which in turn damage molecules leading to aging. Second, the mitochondrial proteome has a manageable number of proteins (1000 to 2000) to study using the current technology in proteomics. Although there are substantial data on the deterioration of mitochondrial function with aging, there is little information on the molecular basis of this phenomenon. We hypothesize that alterations in the mitochondrial proteome have a high probability of playing a major role in the age-related changes in mitochondria function. We proposed to test this hypothesis by performing the first detailed study of global expression and post-translational modifications in mitochondria proteome as a function of age by studying mitochondria isolated from various tissues of mice over their life span and determining the effect of dietary restriction (DR), which retards aging, on the age-related changes observed in the mitochondrial proteome. The Specific Aims of this project are to profile the effects of age and DR on 1) protein expression in the mitochondria proteome, 2) phosphorylation of the mitochondria phosphoproteome, 3) oxidative modification of mitochondria proteins, and 4) mitochondrial function. These studies will be conducted with a series of new proteomics methods we have developed specifically for the examination of protein expression, enzymatic post-translational modifications, and protein oxidation. Recognizing that different proteins are often seen with multidimensional chromatography and gel electrophoresis, we have designed methods that allow us to employ both techniques in our studies of the mitochondrial proteome.