Extended Data Figure 6: Characterization of compound resistant USP7 mutants.
From: Molecular basis of USP7 inhibition by selective small-molecule inhibitors
a–c, SPR-based binding of FT671 (a), FT827 (b) and ubiquitin (c) to indicated USP7 mutants. For compound binding to wild-type USP7CD and Akt-PH controls, see Extended Data Fig. 1b. Sensorgrams are plotted showing two technical replicates. Data are representative of three biological replicates. Values in parentheses represent s.e.m. range. d, Summary table of binding constants observed for experiments in a–c and Extended Data Fig. 1b. Biological repeats are indicated (n = 9 for USP7CD wild type, n = 3 for USP7CD mutants), and values in parentheses indicate s.e.m. e, Circular dichroism profiles of wild-type USP7CD (red), USP7CD(F291N) (orange), USP7CD(Q297A) (green), and USP7CD(Y465N) (blue); each experiment was performed in triplicate. f, Tabulated values for experiments shown in Fig. 3f. g, Structure of activated USP7. The switching loop in USP7 is a point of intrinsic allosteric regulation, provided by a C-terminal module of five ubiquitin-like domains (termed HUBL1 to HUBL5) followed by an activation peptide (amino acids 1084–1102)26 (see Extended Data Fig. 1a). In full-length USP7, the activation peptide stabilizes the active conformation of the switching loop in the ubiquitin-bound state26,27,28, and this can be modulated by USP7-activating proteins, such as GMPS26,29. Shown is the structure of activated USP7 bound to its C-terminal activation peptide. The model was generated from PDB code 5JTV27, and shown are the catalytic domain with ubiquitin in the S1 site (coloured as in Fig. 2a) of molecule 1 in the asymmetric unit and the HUBL5 domain and activation peptide (orange) of molecule 2 in the asymmetric unit, which in the crystal binds to molecule 1 in trans.
