Biophysical Journal 2005-11-01

Fluorescence quenching by TEMPO: a sub-30 A single-molecule ruler.

Peizhi Zhu, Jean-Pierre Clamme, Ashok A Deniz

Index: Biophys. J. 89 , L37-39, (2005)

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Abstract

A series of DNA molecules labeled with 5-carboxytetramethylrhodamine (5-TAMRA) and the small nitroxide radical TEMPO were synthesized and tested to investigate whether the intramolecular quenching efficiency can be used to measure short intramolecular distances in small ensemble and single-molecule experiments. In combination with distance calculations using molecular mechanics modeling, the experimental results from steady-state ensemble fluorescence and fluorescence correlation spectroscopy measurements both show an exponential decrease in the quenching rate constant with the dye-quencher distance in the 10-30 A range. The results demonstrate that TEMPO-5-TAMRA fluorescence quenching is a promising method to measure short distance changes within single biomolecules.