Kellock Jackson, Hopping Gene, Caughey Byron, Daggett Valerie
Department of Bioengineering, University of Washington, Seattle, WA 98105-5013, USA.
Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, MT 59840-2932, USA.
J Mol Biol. 2016 Jun 5;428(11):2317-2328. doi: 10.1016/j.jmb.2016.03.013. Epub 2016 Mar 21.
There is now substantial evidence that soluble oligomers are primary toxic agents in amyloid diseases. The development of an antibody recognizing the toxic soluble oligomeric forms of different and unrelated amyloid species suggests a common conformational intermediate during amyloidogenesis. We previously observed a common occurrence of a novel secondary structure element, which we call α-sheet, in molecular dynamics (MD) simulations of various amyloidogenic proteins, and we hypothesized that the toxic conformer is composed of α-sheet structure. As such, α-sheet may represent a conformational signature of the misfolded intermediates of amyloidogenesis and a potential unique binding target for peptide inhibitors. Recently, we reported the design and characterization of a novel hairpin peptide (α1 or AP90) that adopts stable α-sheet structure and inhibits the aggregation of the β-Amyloid Peptide Aβ42 and transthyretin. AP90 is a 23-residue hairpin peptide featuring alternating D- and L-amino acids with favorable conformational propensities for α-sheet formation, and a designed turn. For this study, we reverse engineered AP90 to identify which of its design features is most responsible for conferring α-sheet stability and inhibitory activity. We present experimental characterization (CD and FTIR) of seven peptides designed to accomplish this. In addition, we measured their ability to inhibit aggregation in three unrelated amyloid species: Aβ42, transthyretin, and human islet amylin polypeptide. We found that a hairpin peptide featuring alternating L- and D-amino acids, independent of sequence, is sufficient for conferring α-sheet structure and inhibition of aggregation. Additionally, we show a correlation between α-sheet structural stability and inhibitory activity.
现在有大量证据表明，可溶性寡聚体是淀粉样疾病中的主要毒性因子。一种能够识别不同且不相关淀粉样物质的毒性可溶性寡聚体形式的抗体的研发，提示了淀粉样蛋白生成过程中存在一种共同的构象中间体。我们之前在各种淀粉样蛋白生成蛋白的分子动力学（MD）模拟中，普遍观察到一种新型二级结构元件的出现，我们将其称为α-片层，并且我们推测毒性构象体是由α-片层结构组成的。因此，α-片层可能代表淀粉样蛋白生成错误折叠中间体的一种构象特征，以及肽类抑制剂潜在的独特结合靶点。最近，我们报道了一种新型发夹肽（α1或AP90）的设计与表征，该肽能形成稳定的α-片层结构，并抑制β-淀粉样肽Aβ42和转甲状腺素蛋白的聚集。AP90是一种由23个残基组成的发夹肽，具有交替的D型和L型氨基酸，具有有利于形成α-片层的构象倾向以及一个设计的转角。在本研究中，我们对AP90进行反向工程，以确定其哪些设计特征对赋予α-片层稳定性和抑制活性最为关键。我们展示了为实现这一目的而设计的七种肽的实验表征（圆二色光谱和傅里叶变换红外光谱）。此外，我们测定了它们抑制三种不相关淀粉样物质聚集的能力：Aβ42、转甲状腺素蛋白和人胰岛淀粉样多肽。我们发现，一种具有交替L型和D型氨基酸的发夹肽，无论其序列如何，都足以赋予α-片层结构并抑制聚集作用。此外，我们还展示了α-片层结构稳定性与抑制活性之间的相关性。
