Hilbich C, Kisters-Woike B, Reed J, Masters C L, Beyreuther K
Center for Molecular Biology, University of Heidelberg, F.R.G.
J Mol Biol. 1991 Mar 5;218(1):149-63. doi: 10.1016/0022-2836(91)90881-6.
The deposition of amyloid beta A4 in the brain is a major pathological hallmark of Alzheimer's disease. Amyloid beta A4 is a peptide composed of 42 or 43 amino acid residues. In brain, it appears in the form of highly insoluble, filamentous aggregates. Using synthetic peptides corresponding to the natural beta A4 sequence as well as analog peptides, we demonstrate requirements for filament formation in vitro. We also determine aggregational properties and the secondary structure of beta A4. A comparison of amino-terminally truncated beta A4 peptides identifies a peptide spanning residues 10 to 43 as a prototype for amyloid beta A4. Infrared spectroscopy of beta A4 peptides in the solid state shows that their secondary structure consists of a beta-turn flanked by two strands of antiparallel beta-pleated sheet. Analog peptides containing a disulfide bridge were designed to stabilize different putative beta-turn positions. Limited proteolysis of these analogs allowed a localization of the central beta-turn at residues 26 to 29 of the entire sequence. Purified beta A4 peptides are soluble in water. Size-exclusion chromatography shows that they form dimers that, according to circular dichroism spectroscopy, adopt a beta-sheet conformation. Upon addition of salts, the bulk fraction of peptides precipitates and adopts a beta-sheet structure. Only a small fraction of peptides remains solubilized. They are monomeric and adopt a random coil conformation. This suggests that the formation of aggregates depends upon a hydrophobic effect that leads to intra- and intermolecular interactions between hydrophobic parts of the beta A4 sequence. This model is sustained by the properties of beta A4 analogs in which hydrophobic residues were substituted. These peptides show a markedly increased solubility in salt solutions and have lost the ability to form filaments. In contrast, the substitution of hydrophilic residues leads only to small deviations in the shape of filaments, indicating that hydrophilic residues contribute to the specificity of interactions between beta A4 peptides.
β淀粉样蛋白A4在大脑中的沉积是阿尔茨海默病的主要病理标志。β淀粉样蛋白A4是一种由42或43个氨基酸残基组成的肽。在大脑中，它以高度不溶性的丝状聚集体形式出现。我们使用与天然βA4序列相对应的合成肽以及类似肽，证明了体外形成细丝的条件。我们还确定了βA4的聚集特性和二级结构。对氨基末端截短的βA4肽进行比较，确定了一个跨越第10至43位残基的肽作为β淀粉样蛋白A4的原型。固态βA4肽的红外光谱表明，它们的二级结构由一个β转角两侧各有两条反平行β折叠片链组成。设计了含有二硫键的类似肽来稳定不同的假定β转角位置。对这些类似物进行有限的蛋白酶解，可将中央β转角定位在整个序列的第26至29位残基处。纯化的βA4肽可溶于水。尺寸排阻色谱显示它们形成二聚体，根据圆二色光谱，这些二聚体具有β折叠构象。加入盐后，大部分肽沉淀并形成β折叠结构。只有一小部分肽保持溶解状态。它们是单体的，具有无规卷曲构象。这表明聚集体的形成取决于疏水效应，该效应导致βA4序列疏水部分之间的分子内和分子间相互作用。这个模型由疏水残基被取代的βA4类似物的特性所支持。这些肽在盐溶液中的溶解度显著增加，并且失去了形成细丝的能力。相反，亲水残基的取代仅导致细丝形状的微小偏差，表明亲水残基有助于βA4肽之间相互作用的特异性。
