Description

UbiQ-314 is an activity-based probe that is based on a ubiquitin variant designed to selectively target USP7. It contains a C-terminal electrophilic propargyl amide (PA, also sometimes abbreviated as Prg). The PA group forms a covalent linkage with the active site Cys residue of USP7 that can be cleaved by acid treatment (5% aq. TFA), allowing for proteomic analyses. The N-terminus is functionalised with a 5-carboxyrhodamine110 dye (cRh110, λex = 480 nm; λem = 520 nm) allowing for a sensitive and fast (in-gel fluorescence) detection. A Cys residue is introduced between the dye and N-terminus for further modification with thiol-reactive tags. UbiQ-314 is based on Ub with the following mutations: T7D, L8Y, I13R, E34L, Q40N, L69A, and L71A. Residue Lys58 contains a biotin tag which besides detection and binding also serves to increase USP7 selectivity by prohibiting binding of most USPs.