Assembly and iron-binding properties of human frataxin, the protein deficient in Friedreich ataxia
Assembly and iron-binding properties of human frataxin, the protein deficient in Friedreich ataxia
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DOI：
10.1093/hmg/11.3.217
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发表时间：
2002-02-01
影响因子：
3.5
通讯作者：
Isaya, G
中科院分区：
文献类型：
--
作者：
Cavadini, P;O'Neill, HA;Isaya, G
弗里德赖希共济失调（FRDA）是一种常染色体隐性退行性疾病，由fraataxin缺乏引起，fraataxin是一种功能未知的保守线粒体蛋白。线粒体铁积累、含铁硫团的酶丢失和氧化损伤增加发生在酵母和小鼠FRDA患者的组织和细胞系中，表明frataxin可能参与了线粒体铁的输出、铁硫团的合成和/或保护免受氧化损伤。我们之前已经证明酵母卵黄蛋白具有铁储存蛋白的结构和功能特征。在本研究中，我们研究了人曲霉素在大肠杆菌和酿酒酵母中的作用。当在大肠杆菌中表达时，成熟形式的人卵黄蛋白组装成稳定的均聚物，每个卵黄蛋白分子可以结合大约10个铁原子。铁负载的均聚物可以通过蛋白质或铁染色在非变性凝胶上检测到，这表明fraataxin和铁之间存在稳定的联系。通过凝胶过滤和电子显微镜分析，均聚物由类似于MDa的球状颗粒和这些颗粒的有序棒状聚合物组成，这些颗粒积聚了小的电子致密核。当人卵黄蛋白前体在酿酒酵母中表达时，由线粒体产生的成熟形式通过凝胶过滤分离成单体和一个高分子量池，分子量约为100 ~ 600 kDa。在小鼠心脏中也存在一个高分子量的纤维蛋白池，这表明纤维蛋白可以在自然条件下组装。在放射性标记的酵母细胞中，人卵黄蛋白通过免疫沉淀回收，每个分子约有5个铁-55原子结合。这些研究结果表明，FRDA是由于frataxin缺乏导致线粒体铁储存减少，这可能损害铁代谢，促进氧化损伤，导致铁的进行性积累。
Friedreich ataxia (FRDA) is an autosomal recessive degenerative disease caused by a deficiency of frataxin, a conserved mitochondrial protein of unknown function. Mitochondrial iron accumulation, loss of iron-sulfur cluster-containing enzymes and increased oxidative damage occur in yeast and mouse frataxin-depleted mutants as well as tissues and cell lines from FRDA patients, suggesting that frataxin may be involved in export of iron from the mitochondria, synthesis of iron-sulfur clusters and/or protection from oxidative damage. We have previously shown that yeast frataxin has structural and functional features of an iron storage protein. In this study we have investigated the function of human frataxin in Escherichia coli and Saccharomyces cerevisiae. When expressed in E.coli, the mature form of human frataxin assembles into a stable homopolymer that can bind approximately 10 atoms of iron per molecule of frataxin. The iron-loaded homopolymer can be detected on non-denaturing gels by either protein or iron staining demonstrating a stable association between frataxin and iron. As analyzed by gel filtration and electron microscopy, the homopolymer consists of globular particles of similar to1 MDa and ordered rod-shaped polymers of these particles that accumulate small electron-dense cores. When the human frataxin precursor is expressed in S.cerevisiae, the mitochondrially generated mature form is separated by gel filtration into monomer and a high molecular weight pool of >600 kDa. A high molecular weight pool of frataxin is also present in mouse heart indicating that frataxin can assemble under native conditions. In radiolabeled yeast cells, human frataxin is recovered by immunoprecipitation with approximately five atoms of Fe-55 bound per molecule. These findings suggest that FRDA results from decreased mitochondrial iron storage due to frataxin deficiency which may impair iron metabolism, promote oxidative damage and lead to progressive iron accumulation.