PROTEOMIC ANALYSIS OF BIOMARKERS AND MECHANISMS OF TOXIC METAL STRESS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 在这个项目中，我们正在使用质谱学蛋白质组学方法来确定汞(II)和有机汞暴露的细胞效应。最初的项目包括对大肠杆菌细胞的研究，这些细胞是否含有携带编码细菌汞解毒途径的多聚操纵子的质粒。我们假设，暴露在汞化合物中可能会导致两种类型的蛋白质修饰，(1)与蛋白质硫醇形成Hg(II)和RHG(I)加合物，以及(2)如果汞相互作用的初始目标刺激活性氧物种的形成，可能导致的氧化修饰。在我的实验室中，工作的一个相当简单的方面是使用基于结构的方法来建立预期的汞(II)-二硫醇和RHG(I)-硫醇蛋白质络合物的预测，以增强用于识别在质谱分析中发现的修饰多肽的数据库。已被证明更重要的第二个方面是开发软件工具，使质谱学数据的分析自动化，以寻找含汞加合物并识别多肽/蛋白质。虽然第二个方面实际上不需要使用嵌合体，但第一个方面需要，预计对识别的目标的进一步分析将涉及蛋白质结构或同源模型的可视化。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. In this project, we are using a mass spectrometry proteomics approach to identify the cellular effects of exposure to Hg(II) and organomercurials. The initial project involves studies of E. coli cells that do or do not harbor a plasmid that carries the mer operon which encodes a bacterial mercury detoxification pathway. We have hypothesized that there may be two types of protein modifications resulting from exposure to the Hg compounds, (1) formation of Hg(II) and RHg(I) adducts with protein thiols, and (2) oxidiative modifications that may result if initial targets of Hg interaction stimulate formation of reactive oxygen species. One fairly straightforward aspect of work in my lab is to use a structure-based approach to build predictions of expected Hg(II)-dithiol and RHg(I)-thiol complexes of proteins to augment the databases used to identify modified peptides found in the mass spec analyses. The second aspect that has proved more important is to develop software tools to automate the analysis of the mass spec data to look for Hg-containing adducts and identify the peptides/proteins. While the second aspect does not really require use of Chimera, the first aspect does and further analysis of identified targets is anticipated to involve visualization of protein structures or homology models.