Conformational transitions of islet amyloid polypeptide (IAPP) in amyloid formation in vitro
Conformational transitions of islet amyloid polypeptide (IAPP) in amyloid formation in vitro
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DOI：
10.1006/jmbi.1999.2646
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发表时间：
1999-04-09
影响因子：
5.6
通讯作者：
Kapurniotu, A
中科院分区：
文献类型：
--
作者：
Kayed, R;Bernhagen, J;Kapurniotu, A
淀粉样蛋白聚集体已被认为是几种致命疾病的病理标志，包括阿尔茨海默病、朊病毒相关疾病和II型糖尿病。胰腺淀粉样变性以胰岛淀粉样多肽（IAPP）为主要成分的淀粉样蛋白沉积为特征。我们遵循的步骤之前IAPP不溶和淀粉样蛋白形成在体外使用各种生化方法，包括过滤试验，远和近紫外圆二色性（CD）分光光度法，1-苯胺基-8-萘磺酸（ANS）结合，原子力（AFM）和电子（EM）显微镜。IAPP不溶解和淀粉样蛋白形成遵循与成核依赖性聚合机制一致的动力学。IAPP淀粉样蛋白形成的成核与预先形成的原纤维的痕迹诱导快速的构象转变成P-片，随后聚集成不溶性淀粉样蛋白原纤维。过渡进行通过熔融球状保形状态与大量的二级结构，波动的叔和季芳族相互作用，和强烈的溶剂暴露的疏水补丁的内容。在5 μ M肽的温度变性途径中，我们发现这种状态主要在约45 ℃下聚集，并且通过长时间暴露于该温度而迅速聚集成淀粉样蛋白，或者当进一步加热至65 ℃时熔化成变性但仍具有结构的IAPP。还发现，在25摄氏度下，在4.25 M GdnHCl诱导的平衡变性过程中，45摄氏度下的状态也会聚集在4.25 M GdnHCl中，并且在聚集成淀粉样纤维之前在溶液中稳定几个小时。我们的研究表明，这种淀粉样状态是IAPP的聚集倾向，部分折叠状态的自相关形式。我们建议，这部分折叠的人口和它的自我相关的形式是在一个浓度依赖性的平衡与非淀粉样蛋白IAPP构象，并可能作为早期，可溶性前体的P-折叠和淀粉样蛋白的形成。我们对IAPP淀粉样蛋白体外形成的分子机制的研究结果有助于深入了解胰腺淀粉样变性和其他淀粉样蛋白相关疾病的发病机制和抑制作用。(C)北京：科学出版社.
Amyloid aggregates have been recognized to be a pathological hallmark of several fatal diseases, including Alzheimer's disease, the prion-related diseases, and type II diabetes. Pancreatic amyloidosis is characterized by the deposition of amyloid consisting of islet amyloid polypeptide (IAPP). We followed the steps preceding IAPP insolubilization and amyloid formation in vitro using a variety of biochemical methods, including a filtration assay, far and near-UV circular dichroism (CD) spectropolarimetry, 1-anilino-8-naphthalenesulfonic acid (ANS) binding, and atomic force (AFM) and electron (EM) microscopy. IAPP insolubilization and amyloid formation followed kinetics that were consistent with the nucleation-dependent polymerization mechanism. Nucleation of IAPP amyloid formation with traces of preformed fibrils induced a rapid conformational transition into P-sheets that subsequently aggregated into insoluble amyloid fibrils. Transition proceeded via a molten globule-like conformeric state with large contents of secondary structure, fluctuating tertiary and quaternary aromatic interactions, and strongly solvent-exposed hydrophobic patches. In the temperature denaturation pathway at 5 mu M peptide, we found that this state was mostly populated at about 45 degrees C, and either aggregated rapidly into amyloid by prolonged exposure to this temperature, or melted into denaturated but still structured IAPP, when heated further to 65 degrees C. The state at 45 degrees C was also found to be populated at 4.25 M GdnHCl at 25 degrees C during GdnHCl-induced equilibrium denaturation, and was stable in solution for several hours before aggregating into amyloid fibrils. Our studies suggested that this amyloidogenic state was a self-associated form of an aggregation-prone, partially folded state of IAPP. We propose that this partially folded population and its self-associated forms are in a concentration-dependent equilibrium with a non-amyloidogenic IAPP conformer and may act as early, soluble precursors of P-sheet and amyloid formation. Our findings on the molecular mechanism of IAPP amyloid formation in vitro should assist in gaining insight into the pathogenesis and inhibition of pancreatic amyloidosis and other amyloid-related diseases. (C) 1999 Academic Press.