Tandem mass tag-based thermal proteome profiling for the discovery of drug-protein interactions in cancer cells

Abstract

Summary: 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).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell Biology
• Cancer
• Molecular Biology
• Protein Biochemistry
• Proteomics
• Mass Spectrometry