以微脂體製備具單一結構之乙型類澱粉樣纖維

生理上乙型類澱粉樣（Aβ）胜肽由它的前驅膜蛋白(APP)經由酵素水解而來，在不同的切位會產生不同長度的Aβ片段。其中以40個胺基酸組成的Aβ40佔絕大多數，其次則為Aβ42、Aβ43等不同的片段。由Aβ所形成的類澱粉樣纖維沉積物是阿茲海默症最明顯的病理特徵之一。文獻中，Aβ纖維具有多種不同的分子結構，且不同構型的Aβ纖維所具有的細胞毒性也不盡相同。我們認為在Aβ聚積的過程初期，不同的成核途徑（初成核、次成核等）可能會引導出不同分子結構的Aβ核種，進而生成不同構型的纖維。為了證明這點，我們嘗試在Aβ40聚合初期時，利用物理性的空間限制減緩Aβ40纖維的形成，令Aβ40核種皆來自初成核，而非與纖維有關的次成核途徑。本研究以逆向蒸發法製備包覆Aβ40單體的微脂體，在經過3－4天的培養後，我們使用對Aβ聚合物有專一性的OMAB抗體辨認，證明微脂體中形成了Aβ40的非纖維狀的核種。接著使用引晶的方式令Aβ40單體依照此核種的結構成長為纖維，並以電子顯微鏡、固態核磁共振技術鑑定纖維結構，證實經由微脂體包覆的步驟，能夠使Aβ40纖維具有幾乎單一的分子結構，初步驗證了類澱粉樣蛋白的成長過程中，最終纖維的結構也許於最初核種的形成途徑就已經決定。此外，我們也試圖以微脂體包覆Aβ42單體，並依照上述方式培養成為Aβ42的核種，並以此對Aβ40單體進行引晶實驗，目的是瞭解不同長度片段的Aβ胜肽之間是否在分子結構較為鬆散的聚合中間態可以產生交叉引晶的現象。最終的實驗結果雖然無法說明兩者在中間態的狀態下確實具有交互作用，但仍提供了一個具有潛力的方向，讓未來的研究者作為參考。

關鍵字

阿茲海默症；類澱粉樣纖維；引晶效應

並列摘要

Physiologically beta amyloid peptides (Aβ) are formed by the enzymatic cleavage of the amyloid precursor protein (APP). Depending on the cutting sites, Aβ peptides may contain 40 to 43 residues, where the 40-residue peptide (Aβ40) is the most abundant species. The plagues formed by the fibrillar aggregation of the Aβ peptides are closely associated with the progression of the AD disease. Amyloid fibrils formed by the 40-residue beta amyloid peptides (Aβ40) are highly polymorphic, with molecular structures that depend on the different kinds of nuclei. In this study, we used the method of reverse-phase evaporation to incubate the Aβ peptides in liposomes, where the peptides were either encapsulated within the liposomes or confined to the interliposomal space. From the results of the dot blot assay using the antibody OMAB, which is specific to the oligomeric aggregates of Aβ40, the Aβ40 peptides incubated in liposomes for 3 to 4 days were proven to be on-pathway non-fibrillar aggregates. These Aβ40 aggregates were used to seed the fibrillization of Aβ40 monomers, and the hence obtained fibril structures were shown to be structurally monomorphic on the basis of solid-state NMR measurements. Our results suggest that the structural polymorphism of Aβ40 may be originated from the early aggregation events of beta-amyloid peptides. In addition, we also attempted to prepare the non-fibrillar aggregates of Aβ42 by the same liposomal process to see whether they can seed the fibrillization of Aβ40 peptides. Our aim is to understand whether the nonfibrillar aggregates of Aβ42 could interact with the Aβ40 monomers. Although the preliminary results cannot unequivocally demonstrate the molecular interaction between Aβ42 and Aβ40 peptides, our approach provides a useful method to probe the possible interactions between on-pathway intermediates of different amyloidogenic peptides.