DNA sequence-dependent enhancement of Cy3 fluorescence

Cyanine dyes are extensively used as fluorescent probes in molecular biology, biochemical and biophysical applications. We investigated the fluorescent properties of Cy3 covalently attached to the 5′ terminus of DNA oligonucleotides, and demonstrated that its fluorescence efficiency and lifetime depend strongly on DNA sequence. DNA sequence determines the extent and nature of the interactions between the dye and the DNA bases, which are responsible for the unusual enhancement in fluorescence observed for a large number of oligonucleotides. Results are discussed in terms of a photoisomerization mechanism that deactivates the excited state and thus competes with fluorescence. The efficiency of isomerization decreases when Cy3-DNA interactions prevent rotation around the double bonds, resulting in an increase in the lifetime of the singlet excited state. We have shown that the ability of Cy3 to interact with DNA depends on the flexibility of the oligonucleotide and the presence of purines in the chain.