Thioflavin T-Silent Denaturation Intermediates Support the Main-Chain-Dominated Architecture of Amyloid Fibrils
Thioflavin T-Silent Denaturation Intermediates Support the Main-Chain-Dominated Architecture of Amyloid Fibrils
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DOI：
10.1021/acs.biochem.6b00231
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发表时间：
2016-07-19
期刊：
影响因子：
2.9
通讯作者：
Goto, Yuji
中科院分区：
文献类型：
--
作者：
Noda, Sayaka;So, Masatomo;Goto, Yuji
超声波被认为是诱导淀粉样蛋白纤维自发形成的最有效的刺激方法之一。当我们用淀粉样特异性硫代黄素T(Tht)荧光监测的β(2)-微球蛋白和胰岛素诱导超声依赖的纤颤时，这两种蛋白在突变期增加后都表现出显著的tht荧光减弱。随着超声功率的增大，THT荧光减弱的速度加快，提示这种减弱是由于预成纤维的部分变性所致。变性的可能中间产物保留了淀粉样蛋白的形态、二级结构和种子电位。当纤维被盐酸胍或十二烷基硫酸钠变性时，也观察到类似的变性中间产物。这些变性中间体的存在与淀粉样蛋白纤维的主链主导结构一致。此外，在所进行的三种变性实验中，胰岛素原纤维比β(2)-微球蛋白原纤维更稳定，这表明各种原纤维的相对稳定性与变性方法无关。
Ultrasonication is considered one of the most effective agitations for inducing the spontaneous formation of amyloid fibrils. When we induced the ultrasonication-dependent fibrillation of beta(2)-microglobulin and insulin monitored by amyloid-specific thioflavin T (ThT) fluorescence, both proteins showed a significant decrease in ThT fluorescence after the burst-phase increase. The decrease in ThT fluorescence was accelerated when the ultrasonic power was stronger, suggesting that this decrease was caused by the partial denaturation of preformed fibrils. The possible intermediates of denaturation retained amyloid-like morphologies, secondary structures, and seeding potentials. Similar denaturation intermediates were also observed when fibrils were denatured by-guanidine hydrochloride or sodium dodecyl sulfate. The presence of these denaturation intermediates is consistent with the main-chain-dominated architecture of amyloid fibrils. Moreover, in the three types of denaturation experiments conducted, insulin fibrils were more stable than beta(2)-microglobulin fibrils, suggesting that the relative stability of various fibrils is independent of the method of denaturation.