α-倒捻子素模拟寡肽破坏Aβ分子内盐桥抑制Aβ聚集作用研究

Tripeptide Mimicing α-Mangostin to Inhibit the Aggregation of Amyloid-β by Breaking the Salt-bridge

关键词:  Aβ聚集抑制剂  α-倒捻子素  模拟肽  噬菌体展示  光学表面等离子共振技术

Key words:  Aβ aggregation inhibitor  α-mangostin  mimic peptide  phage-display  surface plasmon resonance

摘要:

目的 根据特异性结合β淀粉样蛋白（β-amyloid，Aβ）的α-倒捻子素，筛选发现具有抑制Aβ聚集能力的模拟寡肽，阐明其作用机制。方法 通过噬菌体展示技术筛选具有竞争α-倒捻子素与Aβ特异性结合的模拟肽，分析同源序列；根据特征同源序列，合成寡肽；采用光学表面等离子共振技术检测合成寡肽与Aβ的结合力；用特异性检测Aβ聚集程度的荧光素硫磺素T，检测合成寡肽抑制Aβ聚集的能力；采用分子对接技术，对Aβ进行定点突变，探讨合成寡肽抑制Aβ聚集的机制。结果 噬菌体展示技术筛选获得特征同源序列DPN（N为酸性或碱性氨基酸）；根据此特征同源序列合成的寡肽，能特异性结合Aβ；其中DPH能显著性的抑制Aβ的聚集，其作用机制与破坏Aβ分子内第23位天冬氨酸（D23）与第28位赖氨酸（K28）之间的分子内盐桥有关。结论 具有特征性序列DPN的寡肽能竞争α-倒捻子素与Aβ的特异性结合，其中DPH可通过破坏Aβ分子内D23与K28之间的分子内盐桥，显著性的抑制Aβ的聚集。

Abstract:

OBJECTIVE to inhibit the Aβ aggregation and clarify the mechanism by the mimic peptide of α-mangostin which is discovered by binding on β-amyloid (Aβ) specifically. METHODS Phage-display was used to screen the peptides which binded on Aβ specificly against α-mangostin. The homologous sequence achieved by phage-display was analyzed to synthesise the tripeptides of DPN. The binding capability of DPN on Aβ was detected by surface plasmon resonance. The inhibition of DPN to Aβ aggregation by Thioflavin T. Molecular docking and site-specific mutagenesis on Aβ were used to clarify the mechanism. RESULTS The homologous sequence of DPN, where N was an acid or alkaline amino acid, was achieved by the experiment of phage-display. The pepetides with the sequence of DPN, were able to bind Aβ specifically. Especially, DPH would inhibit Aβ aggregation remarkably, which would be induced by the broken of salt-bridge between the residues of Asp23 (D₂₃) and Lys28 (K₂₈). CONCLUSION Pepetides with the sequence of DPN are able to bind Aβ specifically by mimicing α-mangostin. Especially, DPH would inhibit Aβ aggregation remarkably by breaking the salt-bridge between the residues of Asp23 (D₂₃) and Lys28 (K₂₈).