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Abstract：
Mutations in intrinsically disordered proteins drive the irreversible formation of pathological aggregates, a hallmark of neurodegenerative diseases. Here, we present a protocol to pull down fluorescently tagged proteins to characterize their basal oligomeric states. We describe steps for transfection and cell lysis, single-molecule slide preparation and pull-down, and oligomer dissolution. This protocol enables visualization of protein oligomers with single-molecule resolution. In addition, differences in oligomerization may provide insight on condensation or aggregation propensity in differing mutated or cell stress conditions. For complete details on the use and execution of this protocol, please refer to Djaja et al.1.
摘要：
内在无序蛋白质的突变驱动病理性聚集体的不可逆形成，神经退行性疾病的标志。这里,我们提出了一种方案，以拉下荧光标记的蛋白质来表征其基础寡聚状态。我们描述了转染和细胞裂解的步骤，单分子载玻片制备和下拉，和低聚物溶解。该方案能够使单分子分辨率的蛋白质寡聚体可视化。此外,寡聚化的差异可以提供在不同突变或细胞应激条件下缩合或聚集倾向的见解。有关此协议的使用和执行的完整详细信息，PleaserefertoDjajaetal.1.
