CDK5/P35 PHOSPHORYLATION

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 细胞周期蛋白依赖性激酶5(CDK5)是一种调节中枢神经系统神经元结构的蛋白激酶。它只有在与监管合作伙伴配对时才有效；这样的合作伙伴之一被称为p35。当受到压力时，神经元被认为会导致Calain介导的p35裂解成更小的形式，即p25。P25水平的升高与阿尔茨海默病有关，也与CDK5活性增加与肌萎缩侧索硬化症有关。我们正在探索CDK5/p35复合体的磷酸化，因为该激酶似乎受其磷酸化状态的调节。通过质谱学分析，体外用三磷酸腺苷孵育的激酶制剂导致至少两个p35位点和七个cdk5位点的磷酸化。在某些情况下，磷酸化的确切位置已经确定，而另一些情况下，目前正在绘制地图。我们还在进行实验，以确定是否观察到了由于另一种与CDK5/p35复合体共同纯化的激酶而导致的自磷酸化或磷酸化。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Cyclin-dependent kinase 5 (cdk5) is a protein kinase that regulates neuronal structure in the central nervous system. It is active only when paired with a regulatory partner; one such partner is known as p35. When stressed, neurons are believed to cause calpain-mediated cleavage of p35 to a smaller form known as p25. Increased levels of p25 are associated with Alzheimer's disease and have also have been linked, along with increased cdk5 activity, to amyotrophic lateral sclerosis. We are exploring phosphorylation of the cdk5/p35 complex since the kinase appears to be regulated by its phosphorylation state. Incubation of kinase preparations in vitro with ATP results in the phosphorylation of at least two sites within p35 and seven sites within cdk5, as analyzed by mass spectrometry. The exact site of phosphorylation has been determined in some cases, while others are currently being mapped. We are also conducting experiments to determine whether autophosphorylation or phosphorylation due to another kinase co-purified with the cdk5/p35 complex is being observed.