Fluorescence resonance energy transfer as a probe for G-quartet formation by a telomeric repeat

The secondary structure of guanine-rich oligodeoxynucleotides has been investigated with fluorescent probes. Intramolecular folding of a telomeric oligonucleotide into a quadruplex led to fluorescence resonance energy transfer (FRET) between a donor (fluorescein) and an acceptor (tetramethylrhodamine) covalently attached to the 5' and 3' ends of the DNA, respectively. Depending on oligonucleotide length, quenching efficiency varied between 0.45 and 0.72 at 20 degrees C. The conjugation of the dyes to the oligonucleotide had a limited, but significant, influence on the thermodynamics of G-quartet formation. Intramolecular folding was demonstrated from the concentration independence of fluorescence resonance energy transfer over a wide concentration range. Folding of the oligonucleotide was confirmed by UV absorption, UV melting, and circular dichroism experiments. The folding of the G-quartet could be followed at concentrations as low as 100 pM. Fluorescence resonance energy transfer can thus be used to reveal the formation of multistranded DNA structures.