Heterogeneity and Turnover of Intermediates during Amyloid-尾 (A尾) Peptide Aggregation Studied by Fluorescence Correlation Spectroscopy
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- 作者：Ann Tiiman ; J眉ri Jarvet ; Astrid Gr盲slund ; Vladana Vukojevi膰
- 刊名：Biochemistry
- 出版年：2015
- 出版时间：December 15, 2015
- 年：2015
- 卷：54
- 期：49
- 页码：7203-7211
- 全文大小：632K
- ISSN：1520-4995
文摘
Self-assembly of amyloid 尾 (A尾) peptide molecules into large aggregates is a naturally occurring process driven in aqueous solution by a dynamic interplay between hydrophobic interactions among A尾 molecules, which promote aggregation, and steric and overall electrostatic hindrance, which stifles it. A尾 self-association is entropically unfavorable, as it implies order increase in the system, but under favorable kinetic conditions, the process proceeds at appreciable rates, yielding A尾 aggregates of different sizes and structures. Despite the great relevance and extensive research efforts, detailed kinetic mechanisms underlying A尾 aggregation remain only partially understood. In this study, fluorescence correlation spectroscopy (FCS) and Thioflavin T (ThT) were used to monitor the time dependent growth of structured aggregates and characterize multiple components during the aggregation of A尾 peptides in a heterogeneous aqueous solution. To this aim, we collected data during a relatively large number of observation periods, 30 consecutive measurements lasting 10 s each, at what we consider to be a constant time point in the slow aggregation process. This approach enabled monitoring the formation of nanomolar concentrations of structured amyloid aggregates and demonstrated the changing distribution of amyloid aggregate sizes throughout the aggregation process. We identified aggregates of different sizes with molecular weight from 260 to more than 1 脳 106 kDa and revealed the hitherto unobserved kinetic turnover of intermediates during A尾 aggregation. The effect of different A尾 concentrations, A尾:ThT ratios, differences between the 40 (A尾40) and 42 (A尾42) residue long variants of A尾, and the effect of stirring were also examined.