Fig. 2
Characteristics of missense mutations in the human PR-DUB. a Model of the Calypso~Ub–ASX complex generated as described in Fig. 1c, with tumour-derived missense mutations in BAP1 mapped onto the Calypso UCH domain. Residues mutated with low, medium, or high frequency are coloured blue, light orange, or red, respectively. Amino acids located in the active site cleft or at the ubiquitin interface are indicated (numbering refers to the BAP1 sequence; see Supplementary Table 2). The Deubad domain of ASX has been removed for clarity. b Close-up view showing the residues in the NEF-motif of ASX, including Arg288, and the putative interacting amino acids in the modelled ubiquitin (left). Multiple sequence alignment of the residues surrounding the NEF region in ASX, ASXL1, ASXL2, ASXL3, and of the corresponding residues in Rpn13 (right). The NEF-motif is highlighted with a red box; green arrows indicate the residues targeted for mutations. (see Supplementary Fig. 3a and Supplementary Table 3). c Ubiquitin-AMC hydrolysis assays comparing the activity of wild type and mutant Calypso–ASX complexes. Error bars indicate range of measurement (n = 2 independent experiments). DUB, deubiquitinating enzyme; WT, wild type (see Supplementary Fig. 3b, c)
