NEUROFIBROUS PROTEINS IN NORMAL AND AGING BRAIN

这项研究的总体目标是了解核苷酸的作用， 微管相关蛋白(MAP)及其翻译后修饰 在微管介导的细胞功能的调节中。的确有 越来越多的证据表明，协同因素、地图和翻译后 微管蛋白和MAP的修饰显示出年龄相关的变化和一些 这些变化在阿尔茨海默病中可能很重要。具体目标 包括：1.继续进行微管组装的体外探测 GTP和ATP的硫代磷酸类似物和高纯度的无 污染二磷酸核苷酸激酶(NDPK)和ATPase活性 为了加深我们对核苷酸结合机制的理解， 微管蛋白组装过程中的水解性。此外， 微管蛋白刺激的ATPase(MAP-LC)，Kinesin(另一种反应的ATPase 和NDPK将在微管组装系统中进行探测 与核苷酸的硫代磷酸类似物。2.使用微管蛋白酪氨酸 连接酶或胰岛素受体激酶以添加磷酸酪氨酸或 使α-微管蛋白羧基末端的酪氨酸磷酸化。至 分析这一蛋白的磷酸化和去磷酸化的潜在作用 微管蛋白组装中的关键酪氨酸。3.为了进一步描述 微管蛋白和MAP-2对磷酸酪氨酰的作用机制 低分子聚阳离子刺激的磷酸酶活性 蛋白质磷酸酶。这些研究应该有助于阐明 核苷酸、微管相关酶和微管上的磷酸化 细胞骨架和增加我们对微管功能的理解。他们 也应该有助于我们对年龄依赖性改变的理解 大脑中的微管。

The overall aim of this research is to understand the role of nucleotides, microtubule associated proteins (MAPS) and posttranslational modifications in the regulation of microtubule-mediated functions in cells. There is increasing evidence that co-factors, MAPs and posttranslational modifications of tubulin and MAPs show age-dependent changes and some of these changes may be important in Alzheimer's Disease. Specific Aims include: 1. To continue to probe microtubule assembly in vitro using thiophosphate analogues of GTP and ATP and highly purified tubulin free of contaminating nucleotide diphosphate kinase (NDPK) and ATPase activities to further our understanding on the mechanism of nucleotide binding and hydrolysis in tubulin assembly. In addition, the effects of tubulin-stimulated ATPase (MAP-lC), kinesin (another ATPase that reacts with tubulin) and NDPK will be probed in the microtubule assembly system with thiophosphate analogues of nucleotides. 2. To use tubulin tyrosine ligase or insulin receptor kinase to add phosphotyrosine or to phosphorylate tyrosine at the carboxy terminal end of alpha-tubulin. To analyze the potential role of phosphorylation and dephosphorylation of this critical tyrosine in tubulin assembly. 3. To further characterize the mechanism of action of tubulin and MAP-2 regulation on the phosphotyrosyl phosphatase activity of the low-molecular weight polycation-stimulated protein phosphatase. These studies should help to elucidate the role of nucleotides, microtubule associated enzymes and phosphorylation on the cytoskeleton and increase our understanding of microtubule functions. They should also contribute to our understanding of age-dependent alterations of microtubules in brain.