Elucidation of aggregation mechanism of Alzheimer's β-peptides and development of their aggregation inhibitors

阿尔茨海默病的神经病理学特征是淀粉样蛋白在编织实质和皮质血管中的进行性沉积。该沉积主要由40-和42-mer β-淀粉样肽（a - β40, a - β42）组成，具有神经毒性和聚集能力。家族性阿尔茨海默病的脑淀粉样血管病（CAA）与这些a - β肽编码区内的错义突变有关。为了阐明a β肽的聚集机制，我们以高纯度的形式合成了CAA中发现的a β40和a β42的所有变体，并检测了它们对PC12细胞的神经毒性和聚集能力。Aβ40突变体22和23位点的神经毒性均强于野生型Aβ40。22和23位的Aβ42突变体的神经毒性比对应的Aβ40突变体强50 ~ 200倍，这表明这些Aβ42突变体主要参与CAA的发病机制。特别是E…More 22q - a - β42（荷兰）和e22k - a - β42（意大利）广泛聚集，支持了荷兰和意大利患者诊断为遗传性脑出血合并淀粉样变的临床证据。相比之下，a21g （Flemish）突变除了a β突变体的物理化学性质外，还需要其他解释。FI-IR光谱表明，聚集增加了a - β肽的β-薄片含量。然而，转角也是聚集的关键二级结构，因为22和23位的残基更容易形成双残基β-转角，从而显著增强了聚集能力。为了鉴定对β-薄片形成重要的氨基酸残基，我们合成了一系列a - β42的脯氨酸取代突变体，并对它们的聚集能力和神经毒性进行了研究。只有E22P-Aβ42广泛聚集，神经毒性比野生型a β42强，说明15 ~ 21位和24 ~ 32位的残基参与了分子间β片的形成，22和23位的转位形成对a β肽的聚集和神经毒性起关键作用。这些发现为设计新的a - β肽聚集抑制剂奠定了基础。少

Alzheimer's disease is neuropathologically characterized by the progressive deposition of amyloid in the braid parenchyma and cortical blood vessels. This deposition mainly consists of 40-and 42-mer β-amyloid peptides (Aβ40, Aβ42) which show neurotoxicity and aggregative ability. Cerebral amyloid angiopathy (CAA) in familial Alzheimer's disease is related to missense mutations inside the coding region of these Aβpeptides. In order to clarify the aggregation mechanism of Aβpeptides, all of the variants of Aβ40 and Aβ42 found in CAA were synthesized in a highly pure form and examined for neurotoxicity in PC12 cells and aggregative ability.All of the Aβ40 mutants at positions 22 and 23 showed stronger neurotoxicity than wild-type Aβ40. Similar tendency was observed for Aβ42 mutants at positions 22 and 23 whose neurotoxicity was 50〜200 times stronger than that of the corresponding Aβ40 mutants, indicating that these Aβ42 mutants are mainly involved in the pathogenesis of CAA. Especially, E … More 22Q-Aβ42 (Dutch) and E22K-Aβ42 (Italian) aggregated extensively, supporting the clinical evidence that Dutch and Italian patients are diagnosed as hereditary cerebral hemorrhage with amyloidosis. In contrast, A21 G (Flemish) mutation needs alternative explanation with the exception of physicochemical properties of Aβmutants. The FI-IR spectra suggested that aggregation increases theβ-sheet contents of the Aβpeptides. However, turn was also a critical secondary structure for aggregation since residues at positions 22 and 23 that preferably form two-residue β-turn significantly enhanced the aggregative ability.To identify amino acid residues that are important for the β-sheet formation, a series of proline-substituted mutants of Aβ42 was synthesized and their aggregative ability and neurotoxicity were investigated. Only E22P-Aβ42 extensively aggregated with stronger neurotoxicity than wild-type Aβ42, suggesting that the residues at positions 15〜21 and 24〜32 are involved in the intermolecular β-sheet formation, and that turn formation at positions 22 and 23 plays a crucial role in the aggregation and neurotoxicity of Aβpeptides. These findings become a basis for designing new aggregation inhibitors of Aβpeptides. Less