Bioluminescence resonance energy transfer (BRET) is a naturally occurring phenomenon involving the nonradiative energy transfer between a luminescent donor protein (i.e., luciferase) and a fluorescent acceptor protein (e.g., green fluorescent protein).
In the presence of oxygen, luciferase catalyzes the oxidation of its substrate with the concomitant emission of light. When in sufficient physical proximity, the acceptor protein absorbs the energy emitted by the luciferase/substrate reaction and in turn generates a fluorescent signal. The BRET signal is calculated by dividing the acceptor emission intensity by the donor emission intensity:
The magnitude of this ratiometric BRET value is primarily governed by the relative orientation and physical proximity between the donor and acceptor proteins (i.e., the closer the donor and acceptor proteins, the more efficient the energy transfer and thus, the greater the BRET signal). BRET typically occurs in the 1-10 nm range (10-100Å). This spatial range is comparable with the dimensions of biological macromolecules and correlates well with most biological interactions, thus making BRET an ideal system for live-cell monitoring of macromolecular interactions and conformational changes.