2C-BioID: An Advanced Two Component BioID System for Precision Mapping of Protein Interactomes

• Published in: iScience Vol. 10; pp. 40 - 52
• Main Authors: Chojnowski, Alexandre, Sobota, Radoslaw M., Ong, Peh Fern, Xie, Wei, Wong, Xianrong, Dreesen, Oliver, Burke, Brian, Stewart, Colin L.
• Format: Journal Article
• Language: English
• Published: United States Elsevier 21.12.2018
• Subjects: Molecular Biology; Methodology in Biological Sciences; Molecular Interaction
• ISSN: 2589-0042; 2589-0042
• DOI: 10.1016/j.isci.2018.11.023

The modulation of protein-protein interactions (PPIs) is an essential regulatory activity defining diverse cell functions in development and disease. BioID is an unbiased proximity-dependent biotinylation method making use of a biotin-protein ligase fused to a protein of interest and has become an important tool for mapping of PPIs within cellular contexts. We devised an advanced method, 2C-BioID, in which the biotin-protein ligase is kept separate from the protein of interest, until the two are induced to associate by the addition of a dimerizing agent. As proof of principle, we compared the interactomes of lamina-associated polypeptide 2β (LAP2β) with those of lamins A and C, using 2C- and conventional BioID. 2C-BioID greatly enhanced data robustness by facilitating the in silico elimination of non-specific interactors as well as overcoming the problems associated with aberrant protein localization. 2C-BioID therefore significantly strengthens the specificity and reliability of BioID-based interactome analysis, by the more stringent exclusion of false-positives and more efficient intracellular targeting.