Amyloid Protofibrils of Lysozyme Nucleate and Grow Via Oligomer Fusion
Amyloid Protofibrils of Lysozyme Nucleate and Grow Via Oligomer Fusion
复制标题
DOI：
10.1016/j.bpj.2009.01.044
复制
发表时间：
2009-05-06
影响因子：
3.4
通讯作者：
Muschol, Martin
中科院分区：
文献类型：
--
作者：
Hill, Shannon E.;Robinson, Joshua;Muschol, Martin
不溶性淀粉样蛋白原纤维沉积与神经退行性疾病的衰弱性神经元细胞死亡特征之间的联系机制仍然是谜。最近的研究结果表明成熟淀粉样蛋白原纤维的短暂形成的中间体是主要的毒性物质。因此，确定哪些中间聚集体代表通路内前体或通路外侧分支对于理解淀粉样蛋白自组装和设计针对相关毒性物种的治疗方法至关重要。我们用蛋清溶菌酶模型蛋白检测了淀粉样蛋白纤维在酸性溶液中的自组装。结合原位动态光散射和校准原子力显微镜，我们监测了多种瞬态聚集体的成核和生长动力学，并表征了它们的形态和物理尺寸。在高温下孵育后，均匀大小的低聚物以恒定速率形成。经过几个小时的滞后期，原原纤维自发成核。原原纤维的成核动力学及其宽度和高度与低聚物的密切匹配表明原原纤维是通过低聚物融合成核和生长的。原原纤维长到几百纳米后，组装成成熟的原纤维。总的来说，溶菌酶的淀粉样蛋白原纤维组装遵循严格的分层聚集途径，淀粉样蛋白单体、低聚物和原原纤维形成通路上的中间体，依次组装成更复杂的结构。
The mechanisms linking deposits of insoluble amyloid fibrils to the debilitating neuronal cell death characteristic of neurodegenerative diseases remain enigmatic. Recent findings implicate transiently formed intermediates of mature amyloid fibrils as the principal toxic agent. Hence, determining which intermediate aggregates represent on-pathway precursors or off-pathway side branches is critical for understanding amyloid self-assembly, and for devising therapeutic approaches targeting relevant toxic species. We examined amyloid fibril self-assembly in acidic solutions, using the model protein hen egg-white lysozyme. Combining in situ dynamic light scattering with calibrated atomic-force microscopy, we monitored the nucleation and grow:h kinetics of multiple transient aggregate species, and characterized both their morphologies and physical dimensions. Upon incubation at elevated temperatures, uniformly sized oligomers formed at a constant rate. After a lag period of several hours, protofibrils spontaneously nucleated. The nucleation kinetics of protofibrils and the tight match of their widths and heights with those of oligomers imply that protofibrils both nucleated and grew via oligomer fusion. After reaching several hundred nanometers in length, protofibrils assembled into mature fibrils. Overall, the amyloid fibril assembly of lysozyme followed a strict hierarchical aggregation pathway, with amyloid monomers, oligomers, and protofibrils forming on-pathway intermediates for assembly into successively more complex structures.