Template-directed self-assembly and growth of insulin amyloid fibrils
Template-directed self-assembly and growth of insulin amyloid fibrils
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DOI：
10.1002/bit.20486
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发表时间：
2005-06-30
影响因子：
3.8
通讯作者：
Park, CB
中科院分区：
文献类型：
--
作者：
Ha, C;Park, CB
淀粉样蛋白聚集体在组织中的形成是许多神经退行性疾病和II型糖尿病的病理特征。淀粉样蛋白沉积，即单个可溶性淀粉样蛋白分子与预先存在的淀粉样蛋白模板（即斑块）相结合的淀粉样蛋白生长过程，已知对体内淀粉样蛋白的形成至关重要。然而，对天然淀粉样蛋白模板的要求使得淀粉样蛋白沉积研究变得困难和繁琐。在目前的工作中，我们开发了一种新的合成淀粉样蛋白模板，通过将淀粉样蛋白种子共价附着在n -羟基琥珀酰亚胺活化的表面上，其中胰岛素被选为模型淀粉样蛋白。根据非原位原子力显微镜观察，溶液中的胰岛素单体沉积在合成淀粉样蛋白模板上形成原纤维，就像头发生长一样。模板上的纤维形成没有滞后时间，其速度比溶液中快得多。原纤维较长，超过2 μ m，比溶液中的原纤维细得多，这是由于模板表面的成核位点有限和原纤维之间缺乏横向扭转造成的。根据我们使用硫黄素t诱导的荧光、双折射刚果红结合和圆形二色性的研究，在模板上生长的原纤维被鉴定为淀粉样蛋白，通过与模板相互作用后胰岛素单体的构象重排形成。淀粉样蛋白沉积速率随溶液中胰岛素浓度的变化而变化。合成模板上淀粉样蛋白沉积的特征与先前对人类淀粉样斑块进行的研究一致。结果表明，合成的淀粉样蛋白模板可用于体外筛选淀粉样蛋白沉积抑制剂。(c) 2005 Wiley期刊有限公司
The formation of amyloid aggregates in tissue is a pathological feature of many neurodegenerative diseases and type II diabetes. Amyloid deposition, the process of amyloid growth by the association of individual soluble amyloid molecules with a pre-existing amyloid template (i.e., plaque), is known to be critical for amyloid formation in vivo. The requirement for a natural amyloid template, however, has made amyloid deposition study difficult and cumbersome. In the present work, we developed a novel, synthetic amyloid template by attaching amyloid seeds covalently onto an N-hydroxy-succinimide-activated surface, where insulin was chosen as a model amyloidogenic protein. According to ex situ atomic force microscopy observations,insulin monomers in solution were deposited onto the synthetic amyloid template to form fibrils, like hair growth. The fibril formation on the template occurred without lag time, and its rate was highly accelerated than in the solution. The fibrils were long, over 2 mu m, and much thinner than those in the solution, which was caused by limited nucleation sites on the template surface and lack of lateral twisting between fibrils. According to our investigations using thioflavin T-induced fluorescence, birefringent Congo red binding, and circular dichroism, fibrils grown on the template were identified to be amyloids that formed through a conformational rearrangement of insulin monomers upon interaction with the template. The amyloid deposition rate followed saturation kinetics with respect to insulin concentration in the solution. The characteristics of amyloid deposition on the synthetic template were in agreement with previous studies performed with human amyloid plaques. It is demonstrated that the synthetic amyloid template can be used for the screening of inhibitors on amyloid deposition in vitro. (c) 2005 Wiley Periodicals, Inc.