Masino Laura, Nicastro Giuseppe, Menon Rajesh P, Dal Piaz Fabrizio, Calder Lesley, Pastore Annalisa
National Institute for Medical Research, The Ridgeway, London NW7 1AA, UK.
J Mol Biol. 2004 Dec 3;344(4):1021-35. doi: 10.1016/j.jmb.2004.09.065.
Expansion of the polyglutamine (polyQ) region in the protein ataxin-3 is associated with spinocerebellar ataxia type 3, an inherited neurodegenerative disorder that belongs to the family of polyQ diseases. Increasing evidence indicates that protein aggregation and fibre formation play an important role in these pathologies. In a previous study, we determined the domain architecture of ataxin-3, suggesting that it comprises a globular domain, named Josephin, and a more flexible C-terminal region, that includes the polyQ tract. Here, we have characterised for the first time the biophysical properties of the isolated Josephin motif, showing that it is an autonomously folded unit and that it has no significant interactions with the C-terminal region. Study of its thermodynamic stability indicates that Josephin has an intrinsic tendency to aggregate and forms temperature-induced fibrils similar to those described for expanded ataxin-3. We show that, under destabilising conditions, the behaviours of the isolated Josephin domain and ataxin-3 are extremely similar. Our data therefore strongly suggest that the stability and aggregation properties of non-expanded ataxin-3 are determined by those of the Josephin domain, which is sufficient to reproduce the behaviour of the full-length protein. Our data support a mechanism in which the thermodynamic stability of ataxin-3 is governed by the properties of the Josephin domain, but the presence of an expanded polyQ tract increases dramatically the protein's tendency to aggregate.
ataxin-3蛋白中多聚谷氨酰胺（polyQ）区域的扩展与3型脊髓小脑共济失调相关，这是一种遗传性神经退行性疾病，属于polyQ疾病家族。越来越多的证据表明，蛋白质聚集和纤维形成在这些病理过程中起重要作用。在先前的一项研究中，我们确定了ataxin-3的结构域架构，表明它包含一个名为Josephin的球状结构域和一个更灵活的C末端区域，该区域包括polyQ序列。在这里，我们首次表征了分离出的Josephin基序的生物物理特性，表明它是一个自主折叠的单元，并且与C末端区域没有明显的相互作用。对其热力学稳定性的研究表明，Josephin具有内在的聚集倾向，并形成类似于扩展型ataxin-3所描述的温度诱导纤维。我们表明，在不稳定条件下，分离出的Josephin结构域和ataxin-3的行为极其相似。因此，我们的数据强烈表明，未扩展的ataxin-3的稳定性和聚集特性由Josephin结构域的稳定性和聚集特性决定，而Josephin结构域足以重现全长蛋白的行为。我们的数据支持一种机制，即ataxin-3的热力学稳定性由Josephin结构域的特性决定，但扩展的polyQ序列的存在会显著增加蛋白质的聚集倾向。
