Fig. 4: Molecular basis of inhibitor specificity for USP30.
a, Sequence alignment of the indicated human USP DUBs. Arrows indicate the unique Leu328 and Phe453 residues in USP30. b, Close-up view of the compound binding site. c, Superposition with indicated USP DUB structures in complex with inhibitors (PDB 5N9R, 6IIN, 6GH9), highlighting how equivalent Phe and Tyr residues in other human USP DUBs interfere with compound binding. d, Catalytic activities of the indicated wild-type (WT) and mutant USP30 proteins, assessed by Ub–RhoG cleavage. Mean ± s.e.m. (derived from curve fitting; Extended Data Fig. 7). e, Inhibitory potencies of NK036, pre-incubated with the indicated USP30 proteins for 1.5 h, determined from Ub–RhoG cleavage assays. IC50 values are given as mean ± s.e.m. (derived from curve fitting, with activity data for each concentration recorded as n = 3 independent replicates and shown as mean ± s.d.). f, Protein stability of the indicated USP30 proteins in the presence of 20 µM NK036. ΔTm was calculated as Tm of the compound-bound sample subtracted from Tm of the respective apo protein. Mean ± s.d. (n = 3 independent replicates). g, Inhibitory potencies of compound 39, determined as in e. h, Protein stability assessment in the presence of compound 39, determined as in f.
