Figure 2. . Applications of biotinylated reader domain chemical probes.

(A) The general structure of a chemical probe (orange star) tagged with biotin (green circle), with the two moieties connected via a PEG-linker (yellow oval). (B) Biotin tagged probes targeting reader domains are capable of pulling down intact protein complexes as determined by western blot or proteomics analysis. (C) The genomic localization of the probe and its target reader protein can be assessed via Chem-seq which couples chemical affinity capture via a biotinylated ligand with massively parallel DNA sequencing. (D) Biotinylated chemical tools can serve as tracer ligands in in vitro TR-FRET assays to screen for new reader domain ligands and evaluate ligand potency to establish SAR. (E) Protein microarrays can be used to assess chemical probe selectivity within the reader target class by incubation with a biotinylated chemical tool and visualization with fluorescent streptavidin.

FL: Fluorophore; PEG: Polyethylene glycol; SA: Streptavidin; SAR: Structure–activity relationships; TR-FRET: Time-resolved fluorescence energy transfer.