In vitro formation of amyloid from alpha-synuclein is dominated by reactions at hydrophobic interfaces.
In vitro formation of amyloid from alpha-synuclein is dominated by reactions at hydrophobic interfaces.
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DOI：
10.1021/ja102896h
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发表时间：
2010-06
影响因子：
15
通讯作者：
Jeremy Pronchik;Xianglan He;Jason T Giurleo;D. Talaga
中科院分区：
文献类型：
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作者：
Jeremy Pronchik;Xianglan He;Jason T Giurleo;D. Talaga
大多数 α-突触核蛋白 (alphaSyn) 聚集和淀粉样蛋白生成的体外研究都使用空气和/或聚四氟乙烯存在下的搅拌来加速动力学。搅拌的效果隐含或明确地归因于传质或原纤维破碎。本文通过在典型的淀粉样蛋白生成条件下用受控数量的聚四氟乙烯 (PTFE)、聚甲基丙烯酸甲酯 (PMMA) 和硼硅酸盐玻璃制成的球（无顶空）搅拌 alphaSyn 来评估这些假设。使用硫代黄素 T 荧光和原子力显微镜分析淀粉样蛋白。观察到的动力学与 PTFE 表面积成正比；相比之下，PMMA 和玻璃球的影响可以忽略不计。在没有混合球的情况下没有观察到淀粉样蛋白的形成。仅用空气搅拌也显示出加速的动力学，但聚集体形态不同。结果表明，搅拌实验中活跃的机制主要是疏水-水界面的反应。在传质、碎裂和疏水界面假设中，只有最后一个能够解释数据。迄今为止已报道的淀粉样蛋白生成倾向的条件和序列决定因素必须重新解释为反映疏水-水界面的分配。在体内未发现类似的疏水界面。
Most in vitro investigations of alpha-Synuclein (alphaSyn) aggregation and amyloidogenesis use agitation in the presence of air and/or Teflon to accelerate kinetics. The effect of the agitation is implicitly or explicitly attributed to mass transfer or fibril fragmentation. This paper evaluates these hypotheses by agitating alphaSyn under typical amyloidogenic conditions with controlled numbers of balls made of polytetrafluoroethylene (PTFE), polymethylmethacrylate (PMMA), and borosilicate glass with no headspace. Amyloid was assayed using thioflavin T fluorescence and atomic force microscopy. The observed kinetics were proportional to the PTFE surface area; the effects of PMMA and glass balls were negligible by comparison. No amyloid was observed to form in the absence of mixing balls. Agitation with only air also showed accelerated kinetics but different aggregate morphology. The results indicate that the mechanism active in agitation experiments is dominated by reactions at the hydrophobic-water interface. Of the mass transfer, fragmentation, and hydrophobic interface hypotheses, only the last is capable of explaining the data. Condition and sequence determinants of amyloidogenic propensity that have thus far been reported must be reinterpreted as being reflective of partitioning to hydrophobic-water interfaces. Comparable hydrophobic interfaces are not found in vivo.