CAS Key Laboratory of Separation Science for Analytical Chemistry, National Chromatographic R&A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences (CAS), Dalian 116023, China.
University of Chinese Academy of Sciences, Beijing 100049, China.
Molecules. 2023 Jun 20;28(12):4859. doi: 10.3390/molecules28124859.
Although the use of detergents in thermal proteome profiling (TPP) has become a common practice to identify membrane protein targets in complex biological samples, surprisingly, there is no proteome-wide investigation into the impacts of detergent introduction on the target identification performance of TPP. In this study, we assessed the target identification performance of TPP in the presence of a commonly used non-ionic detergent or a zwitterionic detergent using a pan-kinase inhibitor staurosporine, our results showed that the addition of either of these detergents significantly impaired the identification performance of TPP at the optimal temperature for soluble target protein identification. Further investigation showed that detergents destabilized the proteome and increased protein precipitation. By lowering the applied temperature point, the target identification performance of TPP with detergents is significantly improved and is comparable to that in the absence of detergents. Our findings provide valuable insight into how to select the appropriate temperature range when detergents are used in TPP. In addition, our results also suggest that the combination of detergent and heat may serve as a novel precipitation-inducing force that can be applied for target protein identification.
尽管在热蛋白质组学分析（TPP）中使用去污剂已成为识别复杂生物样品中膜蛋白靶标的常见做法，但令人惊讶的是，尚未对去污剂引入对 TPP 靶标识别性能的影响进行全蛋白质组范围的研究。在这项研究中，我们使用泛激酶抑制剂 staurosporine 评估了存在常用非离子型去污剂或两性离子型去污剂时 TPP 的靶标识别性能，结果表明，这两种去污剂中的任何一种都显著降低了可溶性靶蛋白识别的最佳温度下 TPP 的靶标识别性能。进一步的研究表明，去污剂会使蛋白质组不稳定并增加蛋白质沉淀。通过降低应用温度点，TPP 与去污剂的靶标识别性能得到了显著改善，并且可与无去污剂时的性能相媲美。我们的发现为如何在 TPP 中使用去污剂时选择合适的温度范围提供了有价值的见解。此外，我们的结果还表明，去污剂和热的组合可以作为一种新的沉淀诱导力，可用于靶蛋白识别。
