膽固醇在APP蛋白質水解反應中所扮演的角色

Abstract

在阿茲海默氏症(Alzheimer’s disease, AD)的研究領域中，膽固醇與b-amyloid precursor protein (APP)蛋白之間的關係一直是個懸而未決之謎。膽固醇可能借由與APP蛋白有直接的交互作用，而促使APP蛋白走向產生導致阿茲海默氏症的Ab的水解途徑。為了要進一步探討膽固醇與APP蛋白之間的交互作用如何影響APP的水解，我們利用點突變的方式將APP蛋白上與膽固醇做辨認的cholesterol recognition amino acid consensus (CRAC)序列做突變，試著藉由突變產生無法與膽固醇有良好交互作用的APP突變型蛋白。實驗結果發現，APP突變型與lipid raft的標誌蛋白flotillin-1 共分層的程度比野生型來得少；APPY422L與APPS622L在經常用來評估引發致病水解途徑的C99/C83比值上，也較野生型來得低；更進一步的研究發現在去除了膜上的膽固醇後，和僅僅破壞膜上的raft構造，但膜上的膽固醇含量卻仍維持不變時相比，APP蛋白的分層情況並不相同。這些結果指出膽固醇與APP蛋白之間的交互作用力可能造成APP蛋白更容易散布進入lipid raft中，使得APP蛋白更有機會被BACE1水解酶水解，產生Ab片段。

The relationship between cholesterol and b-amyloid precursor protein (APP) processing remains an unsolved mystery in the Alzheimer’s disease (AD) research field. It has been hypothesized that there is a direct interaction between cholesterol and APP, which might facilitate APP being processed into Ab in its pathogenic pathway. To assess the importance of the APP-cholesterol in the regulating of regulating APP processing, we generated several cholesterol-binding-deficient APP mutant proteins via introducing mutations in the domains with the putative cholesterol recognition amino acid consensus (CRAC) sequence of APP. Our results revealed that a smaller portion of APP mutants was co-fractionated with the lipid raft marker flotillin-1 compared to wild-type APP. The C99/C83 ratio, which is usually used in evaluating pathogenesis processing level, is also reduced by Y422L and S622L mutations. Further study shows APP pattern in cholesterol-depleted cells was different from raft-disrupted cells in which the membrane cholesterol content remained unchanged. These data suggested that the interaction between cholesterol and APP could be a driving force for distributing APP into lipid raft, which makes APP more accessible to the pathogenically critical protein BACE1.