Chemoproteomic interrogation of selenocysteine by low-pH isoTOP-ABPP

Selenoproteins comprise a small group of selenocysteine (Sec) containing proteins, often involved in redox homeostasis. While Sec is functionally similar to cysteine (Cys), with both acting as protein-centered nucleophiles, chemoproteomic strategies employing electrophilic probes have often failed to rigorously identify Sec residues, due to their relatively low abundance with respect to Cys across a proteome. To improve the enrichment and detection of selenoproteins, herein we describe a chemoproteomic strategy that relies on the unique properties of Sec as compared to Cys, such as reduced pK_a and the unique isotopic distribution of selenium. Low pH electrophilic probe labeling of mouse proteomes reduces Cys reactivity, resulting in increased identification of most soluble selenoproteins. This quantitative chemoproteomic platform provides a method to reliably measure changes in selenoprotein abundance across growth conditions as well as quantify inhibition by selenoprotein specific inhibitors, such as Auranofin.