Should we fret about FRET?
21 June 2013
Restricted State Selection in Fluorescent Protein Förster Resonance Energy Transfer
Thomas A. Masters, Richard J. Marsh, Daven A. Armoogum, Nick Nicolaou, Banafshé Larijani, Angus J. Bain
When certain molecules get close
together, a few nanometers apart, energy can be transferred between them. If
one molecule is electronically excited, this energy can be transferred to the
other by a process known as Förster resonance energy transfer (FRET). FRET has
been referred to as the “nanometre ruler” and allows the measurement of
intermolecular distances and conformational change without recourse to invasive
techniques such as electron microscopy.
With the development of fluorescent
proteins derived from the Aequorea
victoria jellyfish, it has been possible to induce cells to make these
fluorescent proteins themselves and to attach them to molecules of interest
within the cell itself. This has revolutionised the bio-sciences and led to the
award of the Nobel Prize in Chemistry in 2008.
FRET using fluorescent proteins
is one of the main tools for investigating cell biophysics and cell biochemistry,
two of the most widely used proteins are the enhanced green fluorescent protein
(EGFP) as the donor and the red shifted variant mCherry as the FRET acceptor. However it has been noticed that in some cases, FRET does not work as efficiently as it
should. In a recent collaboration between UCL researchers and Banafshe Larijani’s group at Cancer Research UK’s
London Research Institute, it has been shown that two electronic states in EGFP and mCherry
should be involved in FRET, but in fact only one state in each protein takes
Their work uses the novel combination of fluorescence lifetime and polarisation measurements to reveal this surprising discovery and the results indicate that a number of established FRET techniques would not detect such restrictions. Fretting about FRET is lessened by the time resolved spectroscopic techniques they have developed. This research was supported by Cancer Research UK, EPSRC and UCL Enterprise.