PROTEOMIC ANALYSIS OF BIOMARKERS AND MECHANISMS OF TOXIC METAL STRESS

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 在这个项目中，我们正在使用质谱蛋白质组学的方法来确定暴露于汞（II）和有机汞的细胞效应。 本研究的第一个项目是对E.大肠杆菌细胞中携带或不携带携带编码细菌汞解毒途径的mer操纵子的质粒。 我们假设，可能有两种类型的蛋白质修饰导致暴露于汞化合物，（1）形成汞（II）和RHg（I）与蛋白质硫醇的加合物，（2）氧化修饰，可能导致如果汞相互作用的初始目标刺激活性氧的形成。 在我的实验室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Hg（II）-二硫醇和RHg（I）-硫醇蛋白质复合物的预测，以增加用于识别质谱分析中发现的修饰肽的数据库。 已被证明更重要的第二个方面是开发软件工具来自动分析质谱数据，以寻找含汞加合物并鉴定肽/蛋白质。 虽然第二方面并不真正需要使用嵌合体，但第一方面需要，并且预期对鉴定的靶标的进一步分析涉及蛋白质结构或同源性模型的可视化。

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.