Extended Data Fig. 6: Population profile of PPI probes binding onto unoccupied BCL2 and the cartoon schematic of the experiment.
a-d, PBA binding curves for each PPI probes (eGFP-labeled BIMBH3, BIMEL, BAD, NOXA) to anti-apoptotic proteins (mCherry-labeled BCL2, BCLxL, MCL1) to calculate dissociation constant (Kd). (a) BIMBH3-PBA, (b) BIMEL-PBA, (c) BAD-PBA, (d) NOXA-PBA. e, Comparison of the binding affinities between each binding pairs. f, Correlations between the calculated Kd values and the occupancy values at a fixed PPI probe concentration. g, Dissociation of BCL2-BIM complex after in vitro competition by BAD-eGFP PPI probes (n = 10 independent images). Protein complexes were surface-immobilized by anti-BCL2 IP antibody and detected with anti-BIM detection antibody after the competition. h, Schematic of in vitro PBA competition between BIMBH3-eGFP PPI probe and competitors (PPI probe or BH3 mimetics) on surface-immobilized BCL2 proteins. i, Remained BCL2-BIMBH3 PBA after in vitro binding competition with BH3 mimetics (ABT-199, AZD-5991, WEHI-539). BIMBH3 PPI probe was presented in 10 nM. j,k, BCL2 PBA counts with different PPI probes from four AML cell lines. (j) BCL2-BIMEL PBA, (k) BCL2-BAD PBA (n = 10 independent images). l, Schematic for selective binding of PPI probes for unoccupied BCL2 proteins. Error bars represent means±s.d.
