Fig. 4: InsP6 inhibits Uba6 E1~Ubl thioester formation activity and it enhances its stability.

a, b Kinetic curves of E. coli-derived Uba6 in the presence and absence of InsP6, and insect cell-derived Uba6, in Uba6~Ubl thioester activity assays. Throughout the figure, “Apo” labels refer to E. coli-derived material. Data are presented as mean values +/− SEM (n = 3 technical replicates). c IC50 curves for InsP6 inhibition of the Ub (Left) and FAT10 (Right) thioester formation activities of the indicated Uba6 and Uba1 variants. Data are presented as mean values +/− SEM (n = 3 technical replicates). d Effects of InsP6 and other inositol polyphosphates on the Uba6~Ub (Top) and Uba6~FAT10 (Bottom) thioester formation activities of the indicated Uba6 variants. Assays were conducted in the presence and absence of 100 μM InsPx(3, 5, or 6). Data are presented as mean values +/− SEM (n = 3 technical replicates) and displayed as percentages of the WT value with individual replicates shown as gray circles. e Melting temperatures of the indicated SCCH domain variants as determined by thermal shift assays in the presence and absence of 100 μM InsPx(3, 5, or 6). Data are presented as mean values +/− SEM (n = 3 technical replicates) with individual replicates shown as gray circles. Source data underlying Fig. 4a–e are provided as a Source data file.