A structural approach to understanding the iron-binding properties of phylogenetically different frataxins
A structural approach to understanding the iron-binding properties of phylogenetically different frataxins
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DOI：
10.1093/hmg/11.16.1865
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发表时间：
2002-08-01
影响因子：
3.5
通讯作者：
Pastore, A
中科院分区：
文献类型：
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作者：
Adinolfi, S;Trifuoggi, M;Pastore, A
弗里德赖希共济失调 (FRDA) 是一种常染色体隐性遗传性心脏和神经退行性疾病，是由 frataxin（一种在细胞核中编码的线粒体小蛋白）低表达引起的。在生化水平上，frataxin的缺乏会导致线粒体铁稳态失调和氧化损伤，最终导致神经元死亡。然而，目前尚不清楚 frataxin 是否直接参与铁结合，因为酵母直系同源物（而不是人类蛋白质）已被证明在存在大量铁过量的情况下形成大的聚集体。我们比较了来自 frataxin 家族的三种蛋白质（来自大肠杆菌的细菌 CyaY、酵母 Yfh1 和人类 frataxin）的特性，作为日益复杂的生物体的代表。我们证明这三种蛋白质具有相同的折叠但不同的热稳定性和铁结合特性。虽然人类 frataxin 没有结合铁的倾向，但 CyaY 形成铁促进的聚集体，其行为与酵母 frataxin 类似。然而，聚集可以通过螯合剂或离子强度来竞争。在生理盐条件下，几乎没有观察到聚集。为了识别参与铁促进聚集的蛋白质表面而设计的突变体使我们能够证明该过程是由带负电的表面脊介导的。其中三个残基的突变足以将 CyaY 转化为具有与人 frataxin 相似特性的蛋白质。另一方面，含有大部分保守残基的β片层暴露表面的突变不会影响聚集，这表明铁结合是frataxins更复杂的细胞功能的非保守部分。
Friedreich's ataxia (FRDA), an autosomal recessive cardio- and neurodegenerative disease, is caused by low expression of frataxin, a small mitochondrial protein, encoded in the nucleus. At the biochemical level, the lack of frataxin leads to dysregulation of mitochondrial iron homeostasis and oxidative damage, which eventually causes neuronal death. It is, however, still unclear whether frataxin is directly involved in iron binding, since the yeast orthologue, but not the human protein, has been shown to form large aggregates in the presence of large iron excess. We have compared the properties of three proteins from the frataxin family-the bacterial CyaY from Escherichia coli, the yeast Yfh1 and human frataxin-as representative of organisms of increasing complexity. We show that the three proteins have the same fold but different thermal stabilities and iron-binding properties. While human frataxin has no tendency to bind iron, CyaY forms iron-promoted aggregates with a behaviour similar to that of yeast frataxin. However, aggregation can be competed by chelator agents or by ionic strength. At physiological salt conditions, almost no aggregation is observed. The design of mutants produced to identify the protein surface involved in iron-promoted aggregation allows us to demonstrate that the process is mediated by a negatively charged surface ridge. Mutation of three of these residues is sufficient to convert CyaY in a protein with properties similar to those of human frataxin. On the other hand, mutation of the exposed surface of the beta sheet, which contains most of the conserved residues, does not affect aggregation, suggesting that iron binding is a non-conserved part of a more complex cellular function of frataxins.