Affinity-guided chemical probes for the study of protein interactions

Chemical methods that allow for the targeted labelling of a specific protein within a complex biological environment can enable valuable information regarding the structure and function of proteins to be gained. This thesis explores two different projects where affinity-guided chemical probes were used to study the interactions of proteins, both with small molecules (Chapter 2) and interacting protein partners (Chapter 3). Firstly, chemical labelling methods based on a recognition unit for the protein of interest are reviewed in Chapter 1. Then, Chapter 2 describes how a combination of chemical tools (including photoaffinity, biotinylated and fluorescent probes) were used to study the interaction of a small molecule inducer of human glioblastoma cell death and its relevant target. This work resulted in the identification of HSPD1 as a potential therapeutic target for the treatment of glioblastoma. Chapter 3 details the development of a method for traceless labelling of B-cell lymphoma 2 (BCL-2) family proteins, using a ruthenium-bipyridyl modified peptide. Myeloid cell leukaemia 1 (MCL-1) was rapidly and selectively labelled with fluorescent and biotinylated tags, in vitro, facilitated by the interaction with a peptide mimicking a binding partner. Overall, this thesis demonstrates how affinity-guided labelling of proteins can be used for understanding molecular mechanisms of disease and mapping protein interactomes.