金属离子和酸性pH诱导的β -淀粉样蛋白低聚物不同于中性pH下缓慢自发老化产生的低聚物。

Genevieve M J A Klug, Dusan Losic, Supundi S Subasinghe, Marie-Isabel Aguilar, Lisandra L Martin, David H Small
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引用次数: 101

摘要

用天然和十二烷基硫酸钠/聚丙烯酰胺凝胶电泳检测淀粉样蛋白(Abeta1-40)的聚集和构象,并将结果与原子力显微镜、刚果红结合、沉淀和浊度测定结果进行比较。根据使用的方法,测量的β聚集量是不同的。在pH 5.0或Fe2+, Cu2+或Zn2+存在下孵育15分钟,不改变SDS和天然凝胶上观察到的Abeta低聚物水平。然而,在5天内,Abeta缓慢聚集形成高分子质量的物种被抑制。相比之下,当使用刚刚红结合试验或沉降试验监测Abeta聚集时,在pH 5.0或Fe2+, Cu2+或Zn2+存在下孵育15分钟后,观察到Abeta聚集迅速增加。在浊度测定中测量的低pH, Zn2+或Cu2+诱导的β聚集是可逆的。相比之下,通过原生和SDS/PAGE测量的相当一部分Abeta聚集是稳定的。原子力显微镜研究表明,在pH 5.0或Zn2+存在下,Abeta比pH 7.4时产生更大更松散的棒状聚集体。pH为7.4时老化的Abeta比pH为5.0时老化的Abeta具有更大的细胞毒性。综上所述,结果表明,Abeta寡聚通过两种相互排斥的机制形成两种不同类型的聚集体,这两种聚集体的细胞毒性不同。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Beta-amyloid protein oligomers induced by metal ions and acid pH are distinct from those generated by slow spontaneous ageing at neutral pH.

Amyloid protein (Abeta1-40) aggregation and conformation was examined using native and sodium dodecyl sulfate/polyacrylamide gel electrophoresis, and the results compared with those obtained by atomic force microscopy, and with Congo red binding, sedimentation and turbidity assays. The amount of Abeta aggregation measured was different, depending upon the method used. Incubation for 15 min at pH 5.0 or in the presence of Fe2+, Cu2+ or Zn2+ did not alter the level of Abeta oligomers observed on SDS and native gels. However, the slow aggregation of Abeta to form high molecular mass species over 5 days was inhibited. In contrast, when Abeta aggregation was monitored using a Congo red binding assay or sedimentation assay, a rapid increase in Abeta aggregation was observed after incubation for 15 min at pH 5.0, or in the presence of Fe2+, Cu2+ or Zn2+. The low pH-, Zn2+- or Cu2+-induced Abeta aggregation measured in a turbidity assay was reversible. In contrast, a considerable proportion of the Abeta aggregation measured by native and SDS/PAGE was stable. Atomic force microscopy studies showed that Abeta aged at pH 5.0 or in the presence of Zn2+ produced larger looser rod-shaped aggregates than at pH 7.4. Abeta that had been aged at pH 7.4 was more cytotoxic than Abeta aged at pH 5.0. Taken together, the results suggest that Abeta oligomerizes via two mutually exclusive mechanisms to form two different types of aggregates, which differ in their cytotoxic properties.

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