Core--Metal Ion Regulation /Transport

此核心的目的是为实现此应用程序的目标提供工具和技术支持。该核心将提供对金属的原子吸收分析；对蛋白质进行二维电泳分析的设备和专业知识；用于分析蛋白质相互作用的IAsys仪器；用于分析蛋白质和多肽的质谱学设施；以及用于对细胞和组织中的蛋白质进行免疫定位的共聚焦显微镜设施。原子吸收仪器将允许对铜、铁和硒离子进行灵敏的定量分析，这是研究调节它们的运输和功能的细胞机制所必需的。双向电泳仪将用于分离复杂的细胞蛋白质混合物，以确定信号转导途径的组成部分 调节离子转运，并阐明金属离子毒性的机制。IAsys仪器将提供一种直接的方法来测量离子转运蛋白与细胞中调节其运输和活动的其他成分相互作用的强度和特异性。质谱仪将用于识别离子传输蛋白中的磷酸化位置，识别调节离子传输的新蛋白质，确定转铁蛋白等蛋白质如何相互作用形成蛋白质复合体，并确认将硒半胱氨酸掺入小角度X射线衍射实验制备的蛋白质中的效率。共聚焦显微镜将允许直接显示离子转运蛋白的亚细胞定位，以便了解控制它们在正常和疾病组织中运输的机制。

The purpose of this core is to provide the instrumentation and technical support to fulfill the goals of this application. The core will provide atomic absorption analysis of metals; equipment and expertise to perform two-dimensional electrophoresis analysis of proteins, an IAsys apparatus for the analysis of protein protein interactions; a mass spectrometry facility for the analysis of proteins and peptides; and a confocal microscope facility for the immunolocalization of proteins in cells and tissues. The atomic absorption instrument will allow sensitive quantitative anlysis of copper, iron, and selenium ions necessary to study the cellular mechanisms regulating their transport and function. The two-dimensional electrophoresis equipment will be used to separate complex mixtures of cellular proteins to identify components of signal transduction pathways that regulate ion transport, and to elucidate mechanisms of metal ion toxicity. The IAsys apparatus will provide a direct way to measure the strength and specificity that ion transport proteins interact with other components in the cell that regulate their trafficking and activity. The mass spectrometer will be used to identify phosphorylation sites in ion transport proteins, identify new proteins regulating ion transport, determine how proteins, such as transferrin, interact to form protein complexes, and confirm the efficiency of incorporation of selenocysteine into proteins prepared for small angle x-ray diffraction experiments. The confocal microscope will allow direct visualization of the subcellular localization of ion transport proteins so that the mechanisms controlling their trafficking in normal and diseased tissue can be understood.