Department of Biochemical Engineering, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300350, China.
Key Laboratory of Systems Bioengineering and Frontiers Science Center for Synthetic Biology (Ministry of Education), Tianjin University, Tianjin 300350, China.
ACS Chem Neurosci. 2021 Jan 20;12(2):300-310. doi: 10.1021/acschemneuro.0c00502. Epub 2021 Jan 5.
Cu-mediated amyloid β-protein (Aβ) aggregation is implicated in the pathogenesis of Alzheimer's disease, so it is of significance to understand Cu-mediated conformational transitions of Aβ. Herein, four Aβ mutants were created by using the environment-sensitive cyanophenylalanine to respectively substitute F4, Y10, F19, and F20 residues of Aβ. By using stopped-flow fluorescence spectroscopy and molecular dynamics (MD) simulations, the early stage conformational transitions of the mutants mediated by Cu binding were investigated. The fast kinetics unveils that Cu has more significant influence on the conformational changes of N-terminal (F4 and Y10) than on the central hydrophobic core (CHC, F19, and F20) under different pH conditions (pH 6.6-8.0), especially Y10. Interestingly, lag periods of the conformational transitions are observed for the F19 and F20 mutants at pH 8.0, indicating the slow response of the two mutation sites on the conformational transitions. More importantly, significantly longer lag periods for F20 than for F19 indicate the conduction of the transition from F19 to F20. The conduction time (difference in lag period) decreases from 4.5 s at Cu = 0 to undetectable (<1 ms) at Cu = 10 μM. The significant difference in the response time of F19 and F20 and the fast local conformational changes of Y10 imply that the conformational transitions of Aβ start around Y10. MD simulations support the observation of hydrophobicity increase at N-terminal during the conformational transitions of Aβ-Cu. It also reveals that Y10 is immediately approached by Cu, supporting the speculation that the starting point of conformational transitions of Aβ is near Y10. The work has provided molecular insight into the early stage conformational transitions of Aβ mediated by Cu.
铜介导的淀粉样β蛋白（Aβ）聚集与阿尔茨海默病的发病机制有关，因此了解 Aβ 与铜介导的构象转变具有重要意义。在此，通过使用环境敏感的氰苯丙氨酸分别取代 Aβ 的 F4、Y10、F19 和 F20 残基，创建了四个 Aβ 突变体。通过使用停流荧光光谱法和分子动力学（MD）模拟，研究了突变体与 Cu 结合介导的早期构象转变。快速动力学揭示，在不同 pH 值条件下（pH 6.6-8.0），Cu 对 N 端（F4 和 Y10）构象变化的影响比中央疏水区（CHC，F19 和 F20）更大，尤其是 Y10。有趣的是，在 pH 8.0 时，F19 和 F20 突变体观察到构象转变的滞后期，表明这两个突变位点对构象转变的反应较慢。更重要的是，F20 的滞后期明显长于 F19，表明从 F19 到 F20 的转变。在 Cu = 0 时，转变时间（滞后期差异）从 4.5 s 缩短到 Cu = 10 μM 时的不可检测（<1 ms）。F19 和 F20 的响应时间的显著差异以及 Y10 的快速局部构象变化表明，Aβ 的构象转变始于 Y10 附近。MD 模拟支持在 Aβ-Cu 的构象转变过程中 N 端疏水性增加的观察结果。它还揭示了 Y10 立即被 Cu 接近，支持 Aβ 构象转变的起点在 Y10 附近的推测。这项工作为铜介导的 Aβ 早期构象转变提供了分子见解。
