DOUBLE BRILLIANCE BETA-ARRESTIN: A BIOSENSOR FOR MONITORING THE ACTIVITY OF RECEPTORS AND SIGNALLING MOLECULES, AND METHOD OF USING SAME

摘要

An intramolecular bioluminescence resonance energy transfer (BRET), biosensor for monitoring receptor activity and signalling cascades is disclosed. The ~double-brilliance~ biosensor sandwiches .beta.-arrestin (.beta.-arr) between Renilla luciferase (Luc) and the yellow fluorescent protein (YFP). .beta.-arr associates with G-protein coupled receptors GPCR following receptor activativation, bringing Luc and YPF into close proximity that favours energy transfer. In addition to providing new insights into the agonist-induced conformational rearrangements of .beta.-arr in living cells, the double-brilliance .beta.-arr offers a universal biosensor for GPCR activation, allowing the study of native receptors in large-scale screening analysis. The activity of other signalling molecules known to interact with .beta. arrestin could also be monitored by double brilliance .beta. arr.