Department of Integrative Oncology, BC Cancer Research Institute, Vancouver, BC, Canada; Canada's Michael Smith Genome Sciences Centre, BC Cancer Research Institute, Vancouver, BC, Canada.
Department of Molecular Oncology, University of British Columbia, Vancouver, BC, Canada; Department of Pathology & Laboratory Medicine, University of British Columbia, Vancouver, BC, Canada.
STAR Protoc. 2023 Mar 17;4(1):102012. doi: 10.1016/j.xpro.2022.102012. Epub 2023 Jan 13.
Identification of effector targets is imperative to the characterization of the mechanisms of action of novel small molecules. Here, we describe steps to identify effector drug-protein interactions in lysates derived from cancer cell lines using a thermal proteome profiling (TPP) protocol. Building on existing TTP approaches, we detail the use of an in-solution trypsin digestion technique to streamline sample preparation, a nonparametric analysis to rank proteins for prioritization, and a follow-up strategy for identifying effector interactors. For complete details on the use and execution of this protocol, please refer to Johnson et al. (2022)..
鉴定效应器靶标对于阐明新型小分子的作用机制至关重要。在这里，我们描述了使用热蛋白质组谱（TPP）方案从癌细胞系的裂解物中鉴定效应药物-蛋白相互作用的步骤。在现有的 TTP 方法的基础上，我们详细介绍了使用溶液内胰蛋白酶消化技术来简化样品制备、非参数分析对蛋白质进行优先级排序，以及后续策略来鉴定效应器相互作用物。有关此方案的使用和执行的完整详细信息，请参阅 Johnson 等人（2022 年）。。
