Profiling multi-body interactions of BCL2 with single-molecule co-immunoprecipitation reveals the molecular mechanism of ABT-199 resistance

Abstract

A capability to characterize protein-protein interactions (PPIs) in a quantitative manner with an increased speed would form a technical basis for accelerating drug discovery targeting the PPI network. We here used the single-molecule pull-down and co-IP platform to examine PPI between BCL2 and BH3-only proteins in crude extract environments. We focused on how the PPI strengths changed with single-point BCL2 mutations found in relapsed chronic lymphocytic leukemia showing ABT-199 resistance, where we took a mix-and-match approach to examine various pairs of baits and preys while titrating their concentrations. This allowed us to examine total 21 PPI reactions and 420 data points, forming a high-resolution large data set of the dissociation constants (Kd) and the drug inhibitory constants (Ki). Our data suggest that the different BCL2 mutants take different routes to acquire resistance to ABT-199, demonstrating how large-scale, quantitative PPI data sets reveal insights into the evolving dynamics of PPI networks.