MITOCHONDRIAL IRON STORAGE

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。在正常的衰老过程中，通过过量的饮食或环境暴露，以及调节细胞内金属流量的蛋白质的遗传错误，金属可以在细胞内积累。线粒体铁超载是几种神经系统和代谢性疾病的共同特征。我们已经证明，在铁-硫簇生物合成存在各种错误的酵母中，积累的铁的形式不同。特别是，在体外被Frataxin隔离的类型的亚铁水合物只在表达Frataxin的酵母中发现。我们现在建议将这项工作扩大到比较两种人类疾病--Friedreich‘s共济失调(FRDA)和铁粒母细胞贫血(SA)中的线粒体铁，并开始解决治疗策略的化学问题。以下问题将使用XAS 1解决。Frataxin的铁氧合酶活性如何有助于铁核组装？铁的K边光谱将被收集用于XANES和EXAFS分析，这些铁核是在Frataxin的铁氧化位上被氨基酸取代后组装起来的。2.Frataxin在体内组装的铁核与体内组装的铁核结构相似吗？在体内组装的Frataxin将在非变性凝胶上分离，并在现场收集K-Edge光谱。3.酵母FRDA模型和人类FRDA模型中积累的线粒体铁是否相同？线粒体将从FRDA成纤维细胞培养物中分离出来，并收集K-EDGE光谱用于XANES分析。4.硒能与Frataxin缺乏的线粒体中的铁形成络合物吗？线粒体将从生长在补硒培养基上的Frataxin缺乏的酵母中分离出来，并在铁和硒的K边收集光谱，用于EXAFS分析。5.SA和FRDA之间的铁矿床有区别吗？线粒体将从SA模型中分离出来，并收集用于XANES分析的铁K边光谱。这项工作将有助于开发更好的治疗策略来治疗这些线粒体铁超载障碍。

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. Metals can accumulate in cellular compartments during the normal aging process, through excessive dietary or environmental exposure and from genetic errors in proteins that regulate metal flux within cells. Mitochondrial iron overload is a common feature of several neurological and metabolic diseases. We have shown that in yeast with various errors in iron-sulfur cluster biosynthesis, the form of iron that accumulates differs. In particular, ferrihydrite of the type sequestered by frataxin in vitro is found only in yeast that express frataxin. We now propose to expand this work to compare mitochondrial iron in two human diseases, Friedreich's ataxia (FRDA) and sideroblastic anemia (SA) and to begin to address the chemistry of treatment strategies. The following questions will be addressed using XAS 1. How does frataxin's ferroxidase activity contribute to iron core assembly? Iron K-edge spectra will be collected for XANES and EXAFS analysis of iron cores assembled after amino acid substitutions to frataxin's ferroxidation site. 2. Does frataxin assemble an iron core in vivo that is structurally similar to that assembled in vivo? Frataxin assembled in vivo will be separated on a non-denaturing gel and K-edge spectra collected in situ. 3. Is mitochondrial iron that accumulates in yeast FRDA models and human FRDA the same? Mitochondria will be isolated from FRDA fibroblast cultures and K-edge spectra collected for XANES analysis. 4. Can selenium form a complex with iron in frataxin deficient mitochondria? Mitochondria will be isolated from frataxin-deficient yeast grown on selenium-supplemented media and spectra collected at the iron and selenium K-edges for EXAFS analysis. 5. Do iron deposits differ between SA and FRDA? Mitochondria will be isolated from an SA model and iron K-edge spectra collected for XANES analysis. This work will contribute to the development of better treatment strategies for these mitochondrial iron overload disorders.