Q.38 Fӧrster Resonance Energy Transfer does NOT depend on the
(A) relative orientation of donor and acceptor.
(B) fluorescence quantum yield of acceptor.
(C) distance between donor and acceptor.
(D) overlap between donor’s emission and acceptor’s absorption spectra.
Introduction to FRET
Förster Resonance Energy Transfer (FRET) is a fundamental technique used to measure molecular distances ranging from 1–10 nanometers. It is widely employed to study protein interactions, enzyme mechanisms, and conformational changes in molecular biology — topics often featured in CSIR NET Life Sciences exams.
This distance-dependent, non-radiative process occurs through dipole–dipole coupling between a donor and an acceptor fluorophore. The efficiency of energy transfer is governed by the Förster distance (R0), which defines the separation at which transfer efficiency is 50%.
FRET Efficiency Factors
The efficiency (E) of FRET is expressed by the equation:
E = 1 / [1 + (r / R0)6]
Here, r represents the distance between the donor and acceptor molecules, and R0 is the Förster distance. The Förster radius depends on several parameters according to:
R06 ∝ QD κ2 J / n4
where:
- QD = Fluorescence quantum yield of the donor
- κ2 = Orientation factor between donor and acceptor dipoles (range 0–4)
- J = Spectral overlap integral between donor emission and acceptor absorption
- n = Refractive index of the medium
Therefore, FRET depends critically on donor quantum yield, dipole orientation, and spectral overlap, but not on the fluorescence quantum yield of the acceptor.
Option Analysis
Below is the detailed analysis of each option related to Förster resonance energy transfer dependence:
- (A) Relative orientation of donor and acceptor: FRET efficiency depends on this via the orientation factor (κ2), which influences R0 and dipole–dipole coupling strength.
- (B) Fluorescence quantum yield of acceptor: FRET does not depend on the acceptor’s quantum yield since the process occurs before acceptor emission, involving only donor energy transfer. This is the correct answer.
- (C) Distance between donor and acceptor: The efficiency decreases steeply with distance, following a 1/r6 dependence, making FRET a powerful molecular ruler for nanoscale distances.
- (D) Overlap between donor’s emission and acceptor’s absorption spectra: Essential for resonance energy transfer, quantified by the overlap integral (J).
FRET Dependence Summary
| Factor | Dependence on FRET | Equation Role |
|---|---|---|
| Donor-acceptor distance | Yes, critical | r-6 |
| Spectral overlap | Yes | J in R0 |
| Orientation (κ2) | Yes | Scales R0 |
| Acceptor quantum yield | No | Absent from R0 |
Applications in Biology
FRET plays a vital role in visualizing biomolecular interactions such as protein–protein binding, DNA hybridization, and enzyme activity monitoring. It provides nanometer-scale spatial information essential for understanding molecular mechanisms in biotechnology and cell biology.
Optimum donor–acceptor pairs, for example cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP), achieve a Förster distance (R0) of approximately 5 nm, making them highly efficient for cellular and molecular imaging experiments.
