Pseudophosphorylation of tau protein alters its ability for self-aggregation
Pseudophosphorylation of tau protein alters its ability for self-aggregation
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DOI：
10.1046/j.1471-4159.2003.02287.x
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发表时间：
2004-03-01
影响因子：
4.7
通讯作者：
Holzer, M
中科院分区：
文献类型：
--
作者：
Haase, C;Stieler, JT;Holzer, M
丝状tau蛋白沉积是一组神经退行性疾病（tau病）的病理标志。这些聚集体中的Tau蛋白在不同的磷酸化位点高度磷酸化。尽管肝素诱导的未磷酸化重组tau蛋白聚集可以形成tau蛋白细丝，但尚不清楚tau蛋白磷酸化如何调节聚集行为。由于蛋白激酶的低特异性和体内磷酸化的高度动态转换，在特定的单个或多个位点上分析tau磷酸化的影响受到阻碍。为了克服这个问题，我们采用定点诱变将丝氨酸和苏氨酸转化为天冬氨酸或谷氨酸，从而引入负电荷和类似磷酸化的构象变化。我们测试了14种不同的突变tau蛋白的自聚集倾向和tau蛋白细丝的形成。在肝素、Al3+、Fe2+和Fe3+等不同诱导剂存在下，用硫黄素S荧光监测Tau聚集。我们发现，n端至氨基酸208的突变主要抑制tau聚集，而c端区域的突变主要导致聚集增强。tau中间部分的突变表现出抑制和增强聚集的混合画面。单个氨基酸变化Ser422Glu对所有四种诱导剂都具有有利于聚集的性质。
Filamentous tau protein deposits are a pathological hallmark of a group of neurodegenerative disorders (tauopathies). Tau protein in these aggregates is highly phosphorylated at different phosphorylation sites. Although tau filaments can be formed by heparin-induced aggregation of unphosphorylated recombinant tau, it is not known how tau phosphorylation modulates aggregation behaviour. Analysis of the effect of tau phosphorylation at defined single or multiple sites is hampered by the low specificity of protein kinases and the highly dynamic turnover of phosphorylation in vivo. To overcome this problem we employed site-directed mutagenesis to convert serine and threonine to aspartic acid or glutamic acid, which introduce a negative charge and conformational change that mimic phosphorylation. We tested 14 different mutated tau proteins for their propensity for self-aggregation and formation of tau filaments. Tau aggregation was monitored with thioflavin S fluorescence in the presence of different inducers such as heparin, Al3+, Fe2+ and Fe3+. We found that mutations in the N-terminal portion up to amino acid 208 mainly suppress tau aggregation, whereas mutations in the C-terminal region mainly lead to an enhanced aggregation. Mutations in the middle portion of tau showed a mixed picture of suppression and enhancement of aggregation. A single amino acid change Ser422Glu has aggregation-favouring properties with all four inducers.