XAS OF METALS AND METALLOIDS IN INTRACELLULAR COMPARTMENTS

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。细胞核、线粒体和液泡系统是细胞间室，含有执行特定功能的蛋白质亚群。我们建议使用两种系统来确定金属和类金属的形式。首先，我们建议确定在分离线粒体中积累铁的形式。线粒体铁超载是几种神经和代谢疾病的共同特征。目前，人们假设所有的线粒体铁都是相同的，但没有结构上的证据支持这一观点。铁积聚的形式可能取决于被破坏的途径。为了解决这一问题，我们将利用表征良好的酵母铁/硫簇途径。使用XAS，我们建议表征当与铁/硫簇组装相关的特定蛋白质发生突变时积累的线粒体铁。我们将收集铁k形边缘进行指纹识别，并在适当的时候检查近边缘和EXAFS。由于这些酵母菌中的一些基因具有引起特定疾病的人类同源物，因此对现有物种的明确描述可能导致“定制铁螯合剂”的开发。其次，我们建议在整个动物和器官中确定铜和硒的种类，这些物种已知可以隔离这些化合物。x射线吸收光谱成像是一种相对较新的技术，已被用于询问不同化学物质的空间分布。我们建议将这一技术扩展到两种动物模型中，即果蝇亲铜细胞中铜的区隔化和水船夫中硒的生物积累。硒和铜的XAS光谱中，亚硒酸盐与硒酸盐化合物和铜与铜化合物的近边最大峰位置相差约7eV。这种转变为化学选择性XAS成像提供了充足的对比度。我们的长期目标是将XAS成像应用于哺乳动物系统。

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. The nucleus, mitochondria and vacuolar system are cellular compartments containing subsets of proteins that carry out specific functions. We propose to determine the form of metals and metalloids localized to compartments using two systems. First, we propose to determine the form of accumulated iron in isolated mitochondria. Mitochondrial iron overload is a common feature of several neurological and metabolic diseases. Currently, it is assumed that all excess mitochondrial iron is the same but there is no structural evidence to support this view. The form of iron that accumulates may depend on the pathway that is disrupted. To address this issue we will utilize the well-characterized yeast iron/sulfur cluster pathway. Using XAS, we propose to characterize the mitochondrial iron that accumulates when specific proteins, linked to iron/sulfur cluster assembly, are mutated. We will collect iron K-edges for fingerprinting and examine both the near edge and the EXAFS when appropriate. Since some of these yeast genes have human homologues that cause specific diseases, a definitive characterization of the species present could lead to the development of 'custom iron chelators'. Secondly, we propose to determine copper and selenium species in whole animals and organs that are known to sequester these compounds. X-ray absorption spectroscopy imaging is a relatively new technique that has been used to interrogate the spatial distribution of different chemical species in situ. We propose to extend this technique to two animal models, the compartmentalization of copper in the cuprophilic cells of the fruit fly and selenium bioaccumulation in the water boatman. Both Se and Cu XAS spectra, have a difference of about 7eV between the near edge maximal peak positions of selenite vs selenate and cupric vs cuprous compounds. This shift provides ample contrast for chemically selective XAS imaging. Our long-term goal is to apply XAS imaging to mammalian systems.