Figure 3. Using BioID to generate a history of PPIs or to identify constituents of subcellular domains.

Cellular expression of a protein of interest (bait) fused to a promiscuous biotin ligase called BioID enables biotinylation (denoted by orange circles with B) of proximate proteins within approximately 10nm (light orange disc). This protein labeling is initiated by the addition of a supraphysiological concentration of biotin and requires many hours to generate substantial labeling. Following cell lysis and protein denaturation, biotinylated proteins are captured by affinity purification for identification by mass spectrometry to reveal a history of protein associations that occurred during the labeling period.