Dröden Jessica, Drescher Malte
Department of Chemistry, Konstanz Research School Chemical Biology, University of Konstanz, 78457 Konstanz, Germany.
Biology (Basel). 2023 Jan 3;12(1):79. doi: 10.3390/biology12010079.
Intrinsically disordered proteins (IDPs) are involved in most crucial cellular processes. However, they lack a well-defined fold hampering the investigation of their structural ensemble and interactions. Suitable biophysical methods able to manage their inherent flexibility and broad conformational ensemble are scarce. Here, we used rapid scan (RS) electron paramagnetic resonance (EPR) spectroscopy to study the intermolecular interactions of the IDP α-synuclein (aS). aS aggregation and fibril deposition is the hallmark of Parkinson's disease, and specific point mutations, among them A30P and A53T, were linked to the early onset of the disease. To understand the pathological processes, research intensively investigates aS aggregation kinetics, which was reported to be accelerated in the presence of ethanol. Conventional techniques fail to capture these fast processes due to their limited time resolution and, thus, lose kinetic information. We have demonstrated that RS EPR spectroscopy is suitable for studying aS aggregation by resolving underlying kinetics and highlighting differences in fibrillization behavior. RS EPR spectroscopy outperforms traditional EPR methods in terms of sensitivity by a factor of 5 in our case while significantly reducing data acquisition time. Thus, we were able to sample short time intervals capturing single events taking place during the aggregation process. Further studies will therefore be able to shed light on biological processes proceeding on fast time scales.
内在无序蛋白（IDP）参与了大多数关键的细胞过程。然而，它们缺乏明确的折叠结构，这阻碍了对其结构整体和相互作用的研究。能够处理其固有灵活性和广泛构象整体的合适生物物理方法很少。在这里，我们使用快速扫描（RS）电子顺磁共振（EPR）光谱来研究IDPα-突触核蛋白（aS）的分子间相互作用。aS聚集和纤维沉积是帕金森病的标志，其中特定的点突变，包括A30P和A53T，与该疾病的早发有关。为了理解病理过程，研究集中调查aS聚集动力学，据报道在乙醇存在下其聚集动力学加速。传统技术由于其有限的时间分辨率而无法捕捉这些快速过程，因此会丢失动力学信息。我们已经证明，RS EPR光谱通过解析潜在动力学并突出纤维化行为的差异，适用于研究aS聚集。在我们的案例中，RS EPR光谱在灵敏度方面比传统EPR方法高出5倍，同时显著减少了数据采集时间。因此，我们能够对短时间间隔进行采样，捕捉聚集过程中发生的单个事件。因此，进一步的研究将能够阐明在快速时间尺度上进行的生物过程。
