Award details

Reference fluorescence fluctuation spectroscopy: a new method to study intramolecular motion in biological molecules

Reference	B16491
Principal Investigator / Supervisor	Professor David Klenerman
Co-Investigators / Co-Supervisors	Professor Sir Shankar Balasubramanian
Institution	University of Cambridge
Department	Chemistry
Funding type	Research
Value (£)	112,824
Status	Completed
Type	Research Grant
Start date	21/05/2002
End date	21/05/2004
Duration	24 months

Abstract

We propose to extend our novel method of FRET fluctuation spectroscopy to dual laser excitation and the use of a reporter and reference fluorophor. The reporter fluorescence is quenched as a result of intramolecular motion by a quencher molecule attached to the biomolecule. It should also be possible to use environmental quenching for these experiments. The reporter fluorescence is ratioed to that of the reference fluorophor removing any contribution of diffusion to the correlation function. Since quenching is a step function, a ratio is obtained that is effectively digital i.e. either 1 or 0. The fluctuations in time of this ratio should allow intramolecular dynamics to be determined with superior S/N. In addition the strict requirements in the position of the fluorophors for FRET are removed. We intend to demonstrate this method on DNA hairpin similar to one we have studied previously. Once the method has been validated it can then be used to study intramolecular motion in enzymes, proteins, DNA and other biological molecules of interest. The method also has great potential as an ultra sensitive analytical method particularly for the detection of low levels of DNA.

Summary

unavailable

Committee	Closed Committee - Biomolecular Sciences (BMS)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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