Yusuf Hamied Department of Chemistry, University of Cambridge, Cambridge, UK.
UK Dementia Research Institute, University of Cambridge, Cambridge, UK.
Nat Chem. 2022 Sep;14(9):1045-1053. doi: 10.1038/s41557-022-00976-3. Epub 2022 Jul 7.
The composition of soluble toxic protein aggregates formed in vivo is currently unknown in neurodegenerative diseases, due to their ultra-low concentration in human biofluids and their high degree of heterogeneity. Here we report a method to capture amyloid-containing aggregates in human biofluids in an unbiased way, a process we name amyloid precipitation. We use a structure-specific chemical dimer, a Y-shaped, bio-inspired small molecule with two capture groups, for amyloid precipitation to increase affinity. Our capture molecule for amyloid precipitation (CAP-1) consists of a derivative of Pittsburgh Compound B (dimer) to target the cross β-sheets of amyloids and a biotin moiety for surface immobilization. By coupling CAP-1 to magnetic beads, we demonstrate that we can target the amyloid structure of all protein aggregates present in human cerebrospinal fluid, isolate them for analysis and then characterize them using single-molecule fluorescence imaging and mass spectrometry. Amyloid precipitation enables unbiased determination of the molecular composition and structural features of the in vivo aggregates formed in neurodegenerative diseases.
由于在神经退行性疾病中，可溶毒性蛋白聚集体在人生物体液中的浓度非常低，且具有高度异质性，其体内形成的可溶性毒性蛋白聚集体的组成成分目前尚不清楚。在这里，我们报告了一种在不偏倚的方式下捕获人生物体液中含淀粉样蛋白聚集体的方法，我们将这个过程命名为淀粉样沉淀。我们使用结构特异性化学二聚体（一种具有两个捕获基团的 Y 形仿生小分子）进行淀粉样沉淀以增加亲和力。我们用于淀粉样沉淀的捕获分子（CAP-1）由匹兹堡化合物 B（二聚体）的衍生物组成，用于靶向淀粉样蛋白的交叉 β-片层，以及一个用于表面固定化的生物素部分。通过将 CAP-1 偶联到磁珠上，我们证明我们可以靶向人脑脊液中存在的所有蛋白质聚集体的淀粉样结构，将它们分离出来进行分析，然后使用单分子荧光成像和质谱法对它们进行表征。淀粉样沉淀能够在不偏倚的情况下确定神经退行性疾病中体内形成的聚集体的分子组成和结构特征。
