Fluorescence resonance energy transfer (FRET)

Definition

Fluorescence resonance energy transfer (FRET) is a photophysical phenomenon where energy is transferred non-radiatively from an excited donor fluorophore to an acceptor fluorophore through dipole-dipole interactions. This process occurs when the donor and acceptor are in close proximity, typically within 1-10 nanometers, allowing researchers to study molecular interactions, conformational changes, and distances at the nanoscale. FRET is highly sensitive to the distance and orientation between the two fluorophores, making it a valuable tool in biophysical studies.

5 Must Know Facts For Your Next Test

1. FRET efficiency is influenced by the distance between donor and acceptor; it decreases rapidly as they move apart beyond 10 nanometers.
2. The FRET technique is often used in live cell imaging to observe protein-protein interactions or conformational changes in real time.
3. A common application of FRET is in biosensors, where specific molecular interactions lead to measurable changes in fluorescence.
4. FRET can be quantified using the ratio of fluorescence intensities from the donor and acceptor or through lifetime measurements of the donor.
5. Different pairs of fluorophores can be selected based on their spectral properties, making FRET highly versatile for various experimental setups.

Review Questions

• How does the distance between donor and acceptor fluorophores affect the efficiency of FRET?
  • The efficiency of FRET is highly dependent on the distance between the donor and acceptor fluorophores. As this distance increases beyond approximately 10 nanometers, the likelihood of energy transfer diminishes significantly. This relationship allows researchers to use FRET as a precise measure of molecular proximity, making it a powerful technique for studying interactions at the nanoscale.
• What role does spectral overlap play in the effectiveness of FRET in experimental settings?
  • Spectral overlap is crucial for effective FRET because it involves the overlap between the emission spectrum of the donor and the absorption spectrum of the acceptor. This overlap ensures that when the donor emits light, there is a significant chance that this light will be absorbed by the acceptor, leading to energy transfer. A strong spectral overlap enhances FRET efficiency and allows researchers to select appropriate fluorophore pairs for their experiments.
• Evaluate how FRET can be applied in live cell imaging to study protein interactions, and what implications this has for understanding cellular processes.
  • FRET applications in live cell imaging allow researchers to observe real-time protein interactions and dynamic cellular processes. By tagging proteins with suitable fluorophores, scientists can monitor changes in fluorescence due to energy transfer as proteins come together or move apart within a cell. This technique provides insights into cellular signaling pathways and mechanisms underlying various biological functions, significantly advancing our understanding of complex cellular interactions and their implications in health and disease.

Related terms

Fluorophore: A fluorescent chemical compound that can re-emit light upon light excitation; they are essential for FRET as both donors and acceptors.

Resonance Energy Transfer: The mechanism by which energy is transferred from an excited molecule to another molecule through non-radiative dipole-dipole coupling.

Spectral Overlap:

The overlap between the emission spectrum of the donor fluorophore and the absorption spectrum of the acceptor fluorophore, crucial for efficient FRET.