Polyfluorophore excimers and exciplexes as FRET donors in DNA.
We describe studies aimed at testing whether oligomeric exciplex and excimer fluorophores conjugated to DNA have the potential to act as donors for energy transfer by the Forster mechanism. Oligodeoxyfluorosides (ODFs) are composed of stacked, electronically interacting fluorophores replacing the bases on a DNA scaffold. The monomer chromophores in the twenty tetramer-length ODFs studied here include pyrene (Y), benzopyrene (B), perylene (E), dimethylaminostilbene (D), and a nonfluorescent spacer (S); these are conjugated in varied combinations at the 3' end of a 14mer DNA probe sequence. In the absence of an acceptor chromophore, many of the ODF-DNAs show broad, unstructured long-wavelength emission peaks characteristic of excimer and exciplex excited states, similar to what has been observed for unconjugated ODFs. Although such delocalized excited states have been widely studied, we know of no prior report of their use in FRET. We tested the ability of the twenty ODFs to donate energy to Cy5 and TAMRA dyes conjugated to a complementary strand of DNA, with these acceptors oriented either at the near or far end of the ODF-conjugated probes. Results showed that a number of the ODF fluorophores exhibited relatively efficient energy transfer characteristic of the Forster mechanism, as judged by drops in donor emission quantum yield and fluorescence lifetime, accompanied by increases in intensity of acceptor emission bands. Excimer/exciplex bands in the donors were selectively quenched while shorter-wavelength monomer emission stayed relatively constant, consistent with the notion that the delocalized excited states, rather than individual fluorophores, are the donors. Interestingly, only specific sequences of ODFs were able to act as donors, while others did not, even though their emission wavelengths were similar. The new FRET donors possess large Stokes shifts, which can be beneficial for multiple applications. In addition, all ODFs can be excited at a single wavelength; thus, ODFs may be candidates as "universal FRET donors", thus allowing multicolor FRET of multiple species to be carried out with one excitation.
Teo
Yin Nah
YN
Department of Chemistry, Stanford University, Stanford, California 94305, USA.
Kool
Eric T
ET
D004247:DNA / Q000737:chemistry*; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry*; D008968:Molecular Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*
D005456:Fluorescent Dyes; D009838:Oligodeoxyribonucleotides; D004247:DNA