Proper calibration of ultrasonic power enabled the quantitative analysis of the ultrasonication-induced amyloid formation process
Proper calibration of ultrasonic power enabled the quantitative analysis of the ultrasonication-induced amyloid formation process
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DOI：
10.1002/pro.755
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发表时间：
2012-01-01
期刊：
影响因子：
8
通讯作者：
Kuwata, Kazuo
中科院分区：
文献类型：
--
作者：
Yamaguchi, Kei-ichi;Matsumoto, Tomoharu;Kuwata, Kazuo
为了阐明超声对淀粉样纤维形成的作用机制，我们采用量热法和碘化钾（KI）氧化法对超声功率进行了定量测定，并在适当校准的超声功率下，研究了超声诱导小鼠朊蛋白（mPrP（23231））淀粉样纤维形成的过程。这些方法表明，在我们的系统中的超声波功率范围从0.3到2.7 W，但完全取决于超声波阶段的位置。有趣的是，发现淀粉样蛋白原纤维的成核时间几乎与超声功率成比例地缩短，这表明核形成的发生概率与超声功率成比例地增加。此外，mPrP（23231）形成两种类型的聚集体：刚性纤维和短纤维与无序聚集体，这取决于超声功率。刚性纤维的成核需要大于1.5W的超声功率。而在大于2.6W的强超声功率下，淀粉样纤维形成较早，但同时纤维发生精细的破碎。因此，在所用条件下（每9分钟施加超声处理30秒），约2.0 W的超声功率将适合于形成刚性mPrP（23231）原纤维。由于超声波已被广泛用于体外扩增羊瘙痒症形式的朊病毒蛋白或其他淀粉样蛋白，因此本文提出的量热法和KI氧化法可能有助于确定有效扩增它们所需的足够的超声波功率。
To elucidate the mechanisms of ultrasonication on the amyloid fibril formation, we quantitatively determined the ultrasonic power using both calorimetry and potassium iodide (KI) oxidation, and under the properly calibrated ultrasonic power, we investigated the ultasonication-induced amyloid formation process of the mouse prion protein (mPrP(23231)). These methods revealed that the ultrasonic power in our system ranged from 0.3 to 2.7 W but entirely dependent on the positions of the ultrasonic stage. Intriguingly, the nucleation time of the amyloid fibrils was found to be shortened almost proportionally to the ultrasonic power, indicating that the probability of the occurrence of nucleus formation increases proportionally to the ultrasonic power. Moreover, mPrP(23231) formed two types of aggregates: rigid fibrils and short fibrils with disordered aggregates, depending on the ultrasonic power. The nucleation of rigid fibrils required an ultrasonic power larger than 1.5 W. While at the strong ultrasonic power larger than 2.6 W, amyloid fibrils were formed early, but simultaneously fine fragmentation of fibrils occurred. Thus, an ultrasonic power of approximately 2.0 W would be suitable for the formation of rigid mPrP(23231) fibrils under the conditions utilized (ultrasonication applied for 30 s every 9 min). As ultrasonication has been widely used to amplify the scrapie form of the prion protein, or other amyloids in vitro, the calorimetry and KI oxidation methods proposed here might help determining the adequate ultrasonic powers necessary to amplify them efficiently.