Structural Evolution of Iowa Mutant β-Amyloid Fibrils from Polymorphic to Homogeneous States under Repeated Seeded Growth
Structural Evolution of Iowa Mutant β-Amyloid Fibrils from Polymorphic to Homogeneous States under Repeated Seeded Growth
复制标题
DOI：
10.1021/ja109679q
复制
发表时间：
2011-03-23
影响因子：
15
通讯作者：
Tycko, Robert
中科院分区：
文献类型：
--
作者：
Qiang, Wei;Yau, Wai-Ming;Tycko, Robert
β-淀粉样蛋白原纤维的结构变化对阿尔茨海默病（AD）中这些原纤维的毒性可能很重要。我们描述了一种重复的接种方案，在40个残基的β-淀粉样蛋白原纤维具有Asp 23-to-Asn或爱荷华州突变（D23 N-A β（1-40））的情况下，该方案从多态性初始状态中选择均匀的原纤维结构。我们使用硫磺素T（ThT）荧光，透射电子显微镜（TEM），和固态核磁共振（NMR）跟踪纤维结构的演变，通过多代在此协议下。数据显示，（i）重复接种选择性地扩增单个D23 N-A β（1-40）原纤维结构，其可以是初始多晶型状态的次要组分;（ii）最终结构对生长条件高度敏感，包括pH、温度和搅拌;（iii）尽管初始状态可包括含有反平行和平行β-折叠的原纤维，最终的结构仅含有平行β-折叠，这表明反平行β-折叠结构是化学和动力学亚稳的。此外，我们的数据表明，ThT荧光增强，这是常用的监测淀粉样蛋白原纤维的形成，强烈变化的结构变化，甚至在原纤维组成的相同的多肽。最后，我们提出了一个简单的数学模型，描述了重复播种下的原纤维样品的结构演变。
Structural variations in beta-amyloid fibrils are potentially important to the toxicity of these fibrils in Alzheimer's disease (AD). We describe a repeated seeding protocol that selects a homogeneous fibril structure from a polymorphic initial state in the case of 40-residue beta-amyloid fibrils with the Asp23-to-Asn, or Iowa, mutation (D23N-A beta(1-40)). We use thioflavin T (ThT) fluorescence, transmission electron microscopy (TEM), and solid-state nuclear magnetic resonance (NMR) to track the evolution of fibril structure through multiple generations under this protocol. The data show that (i) repeated seeding selectively amplifies a single D23N-A beta(1-40) fibril structure that can be a minor component of the initial polymorphic state; (ii) the final structure is highly sensitive to growth conditions, including pH, temperature, and agitation; (iii) although the initial state can include fibrils that contain both antiparallel and parallel beta-sheets, the final structures contain only parallel beta-sheets, suggesting that antiparallel beta-sheet structures are thermodynamically and kinetically metastable. Additionally, our data demonstrate that ThT fluorescence enhancements, which are commonly used to monitor amyloid fibril formation, vary strongly with structural variations, even among fibrils comprised of the same polypeptide. Finally, we present a simple mathematical model that describes the structural evolution of fibril samples under repeated seeding.