title: Monitoring protein-protein interactions in living cells by bioluminescence resonance energy transfer (BRET).
abstract: Bioluminescence resonance energy transfer (BRET) allows monitoring of protein-protein interactions in real time in living cells. One candidate interacting protein is fused to a luminescent energy donor, such as Renilla luciferase, and the other to a fluorescent energy acceptor, such the green fluorescent protein (GFP), and the two are then coexpressed in the same cells. If the two proteins interact, their close proximity allows nonradiative energy transfer (BRET) between the luciferase and the GFP. BRET does not occur if the two proteins are separated by more than 100 A, making the technique ideal for monitoring protein-protein interactions in biological systems. This unit describes the use of BRET to study constitutive and agonist-promoted interactions among signaling molecules, as illustrated by the homodimerization of the CXCR4 receptor and the recruitment of beta-arrestin2 to agonist-activated G-protein-coupled receptors. This noninvasive and homogeneous assay provides a robust and sensitive proteomic platform with applications for basic science research and drug discovery.
authors.affiliation: University of Montreal, Montreal, Quebec, Canada.
authors.forename: Fadi F
authors.initials: FF
authors.lastname: Hamdan
authors.forename: Yann
authors.initials: Y
authors.lastname: Percherancier
authors.forename: Billy
authors.initials: B
authors.lastname: Breton
authors.forename: Michel
authors.initials: M
authors.lastname: Bouvier
mesh_terms: D000818:Animals; D002461:Cell Line, Transformed; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D015870:Gene Expression / Q000502:physiology; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods*; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection / Q000379:methods
chemical_list: D008164:Luminescent Proteins; D011506:Proteins