Sugiki Toshihiko, Yoshiura Chie, Kofuku Yutaka, Ueda Takumi, Shimada Ichio, Takahashi Hideo
Japan Biological Informatics Consortium (JBiC), Aomi, Koto-ku, Tokyo 135-8073, Japan.
Protein Sci. 2009 May;18(5):1115-20. doi: 10.1002/pro.92.
Protein aggregation is an essential molecular event in a wide variety of biological situations, and is a causal factor in several degenerative diseases. The aggregation of proteins also frequently hampers structural biological analyses, such as solution NMR studies. Therefore, precise detection and characterization of protein aggregation are of crucial importance for various research fields. In this study, we demonstrate that fluorescence correlation spectroscopy (FCS) using a single-molecule fluorescence detection system enables the detection of otherwise invisible aggregation of proteins at higher protein concentrations, which are suitable for structural biological experiments, and consumes relatively small amounts of protein over a short measurement time. Furthermore, utilizing FCS, we established a method for high-throughput screening of protein aggregation and optimal solution conditions for structural biological experiments.
蛋白质聚集是多种生物学情况下的一个重要分子事件，也是几种退行性疾病的致病因素。蛋白质聚集还经常妨碍结构生物学分析，如溶液核磁共振研究。因此，精确检测和表征蛋白质聚集对于各个研究领域至关重要。在本研究中，我们证明使用单分子荧光检测系统的荧光相关光谱法（FCS）能够在较高蛋白质浓度下检测到原本不可见的蛋白质聚集，这种浓度适用于结构生物学实验，并且在短测量时间内消耗的蛋白质量相对较少。此外，利用FCS，我们建立了一种高通量筛选蛋白质聚集的方法以及用于结构生物学实验的最佳溶液条件。
