Department of Physics, University of Central Florida , Physical Sciences Bldg., Room 456, 4111 Libra Drive, Orlando, Florida 32816, United States.
J Phys Chem B. 2017 Nov 16;121(45):10293-10305. doi: 10.1021/acs.jpcb.7b07002. Epub 2017 Nov 1.
Amyloid β (Aβ) peptide contributes to Alzheimer's disease by a yet unidentified mechanism. In the brain tissue, Aβ occurs in various forms, including an undecapeptide Aβ, which exerts a neurotoxic effect through the mitochondrial dysfunction and/or Ca-permeable pore formation in cell membranes. This work was aimed at the biophysical characterization of membrane binding and pore formation by Aβ. Interaction of Aβ with anionic and zwitterionic membranes was analyzed by microelectrophoresis. In pore formation experiments, Aβ was incubated in aqueous buffer to form oligomers and added to Quin-2-loaded vesicles. Gradual increase in Quin-2 fluorescence was interpreted in terms of membrane pore formation by the peptide, Ca influx, and binding to intravesicular Quin-2. The kinetics and magnitude of this process were used to evaluate the rate constant of pore formation, peptide-peptide association constants, and the oligomeric state of the pores. Decrease in membrane anionic charge and high ionic strength conditions significantly suppressed membrane binding and pore formation, indicating the importance of electrostatic interactions in these events. Circular dichroism spectroscopy showed that Aβ forms the most efficient pores in β-sheet conformation. The data are consistent with an oligo-oligomeric pore model composed of up to eight peptide units, each containing 6-8 monomers.
淀粉样 β 肽（Aβ）通过一个尚未确定的机制导致阿尔茨海默病。在脑组织中，Aβ 以多种形式存在，包括具有神经毒性作用的十肽 Aβ，其通过线粒体功能障碍和/或细胞膜中 Ca 通透性孔的形成发挥作用。这项工作旨在对 Aβ 的膜结合和孔形成进行生物物理特性分析。通过微电泳分析 Aβ 与阴离子和两性离子膜的相互作用。在孔形成实验中，将 Aβ 在水性缓冲液中孵育以形成低聚物，然后加入负载 Quin-2 的囊泡。肽形成孔时，逐渐增加 Quin-2 荧光，可以解释为 Ca 内流和与囊内 Quin-2 结合，导致膜通透性增加。该过程的动力学和幅度用于评估孔形成的速率常数、肽-肽缔合常数以及孔的寡聚状态。减少膜阴离子电荷和高离子强度条件显著抑制了膜结合和孔形成，表明静电相互作用在这些事件中的重要性。圆二色性光谱表明 Aβ 在 β-折叠构象下形成最有效的孔。这些数据与由多达八个肽单元组成的寡聚孔模型一致，每个单元包含 6-8 个单体。
