In vitro CRBN-BTK dimerization

BODIPY-labeled CRBN in complex with damage-specific DNA-binding protein 1 (DDB1) harboring an internal deletion of the flexible β propeller B domain (DDB1ΔB-CRBN)²² and in vitro–biotinylated BTK were treated with increasing concentrations of the indicated compounds in the presence of tracer amounts of terbium-streptavidin. Compound-induced recruitment of BTK to CRBN was quantified following the 520/490 time-resolved fluorescence resonance energy transfer (FRET) ratio²² using a Pherastar plate reader (BMG) utilizing 2 synchronized photomultiplier tubes to reduce background.