Mechanism of sequence-specific fluorescent detection of DNA by N-methyl-imidazole, N-methyl-pyrrole, and β-alanine linked polyamides

The fluorescence from the tetramethylrhodamine (TMR) moiety in hairpin polyamide-TMR conjugates is quenched in solution, but restored upon sequence- specific binding to doubled- stranded DNA. This fluorescence amplification when bound to the target DNA sequence makes polyamidc-TMR conjugates potentially useful for the detection of specific DNA sequences in homogeneous solution. Time-resolved and steady-state spectroscopic measurements indicate that a ground-state complex forms between the TMR and polyamide functionalities in the absence of DNA. This intramolecular complex likely facilitates electron transfer from the polyamide N-methyl-pyrrole moieties to the TMR excited state, quenching fluorescence. Binding of the polyamide-TMR probe to the target DNA sequence disrupts the TMR-polyamide interaction, resulting in the observed fluorescence increase.