Fig. 4.

IL-loop contacts the catalytic domain of USP25 in the tetramer. a Surface and cartoon representation of interface between the IL-loop domain (green), “kink” region (blue), and the catalytic domain in the tetramer assembly. b Structure overlapping of USP25 and USP7 structures (PDB code 5JTJ), depicting the S1 ubiquitin-binding domain in contact with either ubiquitin in complex with USP7 structure (light blue) or with the IL-loop domain in the USP25 tetramer (red ribbon). c Surface representation of the S1 ubiquitin-binding pocket (green) of the USP25 catalytic domain in complex with the IL-loop insertion with labeled residues depicted in stick representation. d Cartoon representation of the IL-loop binding interface with labeled residues depicted in stick representation (green). IL-loop is depicted as a thin line (red). e Deubiquitinating activity assays with Ub-AMC substrate of either USP25 wild-type and IL-loop point mutants: purified tetramer and dimer, P521S, F522G, S525P, P528G, and P535L (first and second panel); or USP25 wild-type and S1 ubiquitin-binding residues point mutants: E373A, Q322A, L271W, and C651F (third and fourth panel)