FRET from Multiple Pathways in Fluorophore-Labeled DNA

Because of their ease of design and assembly, DNA scaffolds provide a valuable means for organizing fluorophores into complex light harvesting antennae. However, as the size and complexity of the DNA–fluorophore network grows, it can be difficult to fully understand energy transfer properties because of the large number of dipolar interactions between fluorophores. Here, we investigate simple DNA–fluorophore networks that represent elements of the more complex networks and provide insight into the Forster Resonance Energy Transfer (FRET) processes in the presence of multiple pathways. These FRET networks consist of up to two Cy3 donor fluorophores and two Cy3.5 acceptor fluorophores that are linked to a rigid dual-rail DNA scaffold with short interfluorophore separation corresponding to 10 DNA base pairs (∼34 A). This configuration results in five FRET pathways: four hetero-FRET and one homo-FRET pathway. The FRET properties are characterized using a combination of steady-state and time-resolved spectrosc...