Photoaffinity strategies to profile cholesterol metabolite interactomes and binding sites

Cholesterol and its metabolites regulate diverse and essential roles in biological systems and feature prominently in human health and disease. Key to the activity of these molecules are the proteins that they interact with in a given cell type and condition. In this Chapter, we describe the use of chemical photoaffinity probes to map the interactome of cholesterol metabolites throughout the proteome and to pinpoint the site of binding within a protein of interest. We pay close attention to variables that influence the study of cholesterol-based molecules, describing experimental and data analysis parameters to optimize proteome/binding site coverage and robustness of results.