Nuclear magnetic resonance evidence for the dimer formation of beta amyloid peptide 1-42 in 1,1,1,3,3,3-hexafluoro-2-propanol.
Nuclear magnetic resonance evidence for the dimer formation of beta amyloid peptide 1-42 in 1,1,1,3,3,3-hexafluoro-2-propanol.
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核磁共振证据证明 1,1,1,3,3,3-六氟-2-丙醇中 β 淀粉样肽 1-42 形成二聚体。
DOI：
10.1016/j.ab.2015.12.021
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发表时间：
2016
影响因子：
2.9
通讯作者：
Hidekazu Hiroaki
中科院分区：
文献类型：
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作者：
Yoshiki Shigemitsu;Naoko Iwaya;Natsuko Goda;Mizuki Matsuzaki;Takeshi Tenno;Akihiro Narita;Minako Hoshi;Hidekazu Hiroaki
阿尔茨海默病涉及老年斑的积累，其中沉积了淀粉样β蛋白(Aβ)多肽的丝状聚集体。最近的研究表明，Aβ多肽的齐聚途径可能是复杂的。为了更详细地了解Aβ(1-42)齐聚物的形成机理，我们建立了一种适合于核磁共振研究的15N标记Aβ(1-42)的制备方法。对于物理化学研究，起始的蛋白质材料应该是单体的，并且必须去除所有的Aβ聚集体。在这里，我们成功地通过高效液相色谱从易于聚集的组分中分离出Aβ(1-42)的“抗沉淀”组分，甚至从细菌过度表达的Aβ(1-42)中也成功地进行了分离。然而，经1，1，1，3，3，3-六氟-2-丙醇处理后的两个Aβ(1-42)组分都形成了淀粉样纤维。这表明，在HFIP处理过程中，“聚集种子”并没有完全单体。此外，1H-15N核磁共振谱表明，β(1-42)在HFIP中表现为单体-二聚体平衡。在硫代黄素T荧光分析中，我们证明了在HFIP前处理过程中Aβ的初始浓度改变了Aβ纤维形成的动力学轮廓。在研究Aβ(1-42)肽时，这里描述的发现应该能确保结果的重复性。
Alzheimer's disease involves accumulation of senile plaques in which filamentous aggregates of amyloid beta (Aβ) peptides are deposited. Recent studies demonstrate that oligomerization pathways of Aβ peptides may be complicated. To understand the mechanisms of Aβ(1–42) oligomer formation in more detail, we have established a method to produce15N-labeled Aβ(1–42) suited for nuclear magnetic resonance (NMR) studies. For physicochemical studies, the starting protein material should be solely monomeric and all Aβ aggregates must be removed. Here, we succeeded in fractionating a “precipitation-resistant” fraction of Aβ(1–42) from an “aggregation-prone” fraction by high-performance liquid chromatography (HPLC), even from bacterially overexpressed Aβ(1–42). However, both Aβ(1–42) fractions after 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) treatment formed amyloid fibrils. This indicates that the “aggregation seed” was not completely monomerized during HFIP treatment. In addition, Aβ(1–42) dissolved in HFIP was found to display a monomer–dimer equilibrium, as shown by two-dimensional1H–15N NMR. We demonstrated that the initial concentration of Aβ during the HFIP pretreatment altered the kinetic profiles of Aβ fibril formation in a thioflavin T fluorescence assay. The findings described here should ensure reproducible results when studying the Aβ(1–42) peptide.