HUMAN ETHE1: ACTIVITY AND CATALYTIC MECHANISM

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。列出的机构是 研究中心，而研究中心不一定是研究者所在的机构。 微量元素对所有生物体都是必不可少的。同时在细胞中游离金属离子的浓度非常低，因为游离的氧化还原活性金属可以产生自由基，这可能导致细胞损伤。其他的，如Ca和Zn参与信号传导过程。这意味着需要严格监管。在哺乳动物中，调节功能障碍导致严重疾病。细菌在金属含量过高或过低的情况下也会遭受类似的问题。鉴于哺乳动物和细菌的微量元素调控是基于不同的蛋白质网络，细菌金属调节剂作为潜在的药物靶点引起了相当大的关注。在过去的几年里，我们已经建立了我们的研究细菌金属调节。 病原菌如M.结核病必须与铁螯合作斗争，以便在人体内存活。铁代谢是通过控制参与铁吸收、运输和储存的基因的转录来调节的。In M.在结核病中，铁摄取调节因子A（FurA）被Fe 2+激活以特异性结合其靶DNA序列，从而抑制下游基因。这些基因在同源情况下包括毒力因子和细菌铁稳态所需的几种蛋白质。通过X射线吸收光谱，我们将确定蛋白质的金属结合位点。 这允许区分两个结构和功能不同的FurAMtb金属结合位点，并提供了细致的描述加上定性和定量表征。 在这里，我们提出了X射线吸收光谱测量结核分枝杆菌铁吸收调节剂FurA进一步表征这一重要的基因产物的存在和不存在一个新的铅化合物。为了比较，我们将包括Fur_AF从嗜酸氧化亚铁硫杆菌，革兰氏阴性菌生活在pH 2 - 2的高浓度的可溶性亚铁和三价铁。

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. Trace elements are essential for all living organisms. At the same time in cells the concentration of free metal ions is very low, because free redox-active metals can generate radicals, which can lead to cell damage. Others, such as Ca and Zn are participating in signaling processes. This implies the need for tight regulation. In mammals dysfunction of the regulation causes severe diseases. Bacteria would suffer from similar problems in case of too high or too low metal contents. Given the fact that mammalian and bacterial trace element regulation is based on different protein networks, the bacterial metal regulators have attracted considerable attention as potential drug targets. During the past years we have established our research on bacterial metal regulation. Pathogens, such as M. tuberculosis have to contend with iron sequestration in order to survive in the human body. Iron metabolism is regulated by controlling transcription of genes involved in iron uptake, transport and storage. In M. tuberculosis the ferric uptake regulator A (FurA) is activated by Fe2+ to bind specifically to its target DNA sequence thereby repressing the downstream genes. These genes include in homologous cases virulence factors and several proteins required by the bacterium for iron homeostasis. By x-ray absorption spectroscopy we will determine the metal binding sites of the protein. This allows distinguishing between two structurally and functionally distinct FurAMtb metal binding sites and provides a meticulous description plus a qualitative and quantitative characterization of them. Here we propose x-ray absorption spectroscopy measurements on the Mycobacterium tuberculosis iron uptake regulator FurA to further characterize this important gene product in the absence and presence of a new lead compound. For comparison we will include Fur_AF from Acidithiobacillus ferrooxidans, a Gram-negative bacterium living at pH2 2 in high concentrations of soluble ferrous and ferric iron.