Extended Data Fig. 2: Identification of ATP5B as the target of apoptolidin A using comparative proteomics.

a, Gel-based profiling of Apop A-PA targets in MV-4–11 with ± 1 μM Apop A-PA, ± 20 μM Apop A after affinity enrichment using streptavidin resin prior to digestion and TMT labeling imaged by rhodamine fluorescence and after silver staining; b, schematic of the TMT multiplexing strategy for to combine vehicle, probe, and competition, conditions from two separate biological replicates; c, Volcano plot of proteins significantly enriched in ‘Probe’ condition compared to ‘Vehicle’ analyzed by TMT-multiplexed MuDPIT proteomics; d, Volcano plot of proteins significantly enriched in ‘Competition’ condition compared to ‘Vehicle’ analyzed by TMT-multiplexed MuDPIT proteomics; e, Coomassie stained gel-based profiling of Apop A-PA targets in MV-4–11 cells – 50 kDa band from probe and competition conditions were cut and subjected to in-gel digestion (see Supplementary Table 1); f, Immunoblot showing pulldown of FLAG tagged ATP5B expressed in H293 landing pad cells.