PUB - Publikationen an der Universität Bielefeld

A novel picosecond laser approach is used to investigate the intrinsic fluorescence of several oligonucleotides. All biomolecules are excited at 283 nm with laser pulses of typically 80 ps duration and an energy of 250 mu J; concentrations were on the order of 10(-5) M. Detection of the resulting fluorescence behind a spectrometer with a streak camera permits the simultaneous acquisition of spectral and lifetime information in two-dimensional images. In a systematic study, the fluorescence spectra and the associated temporal decays are analyzed with respect to monomer and potential excimer components. For this, the nucleotides AMP, CMP, GMP, and TMP are studied as well as homo-oligonucleotides of the type d(X)(n) with variable sequence length of n = 2-15, enabling a comparison of the emission characteristics of these single-stranded compounds under physiologic conditions in solution at room temperature. Also, the influence of conformational changes on the fluorescence response is investigated using mixtures of complementary oligonucleotides d(X)(15)xd(Y)(15) With the combinations X = A, Y = T and X = G, Y = C. These structures, which form double helices, differ in flexibility and stacking geometry from the single-stranded compounds. From experiments with self-complementary variants with alternating base sequences of the type d(XY)(8) with XY = AT and GC, information on exciplex formation tendencies is obtained for these compounds, which also form double helices in solution. Preliminary results of time-dependent fluorescence anisotropy measurements with this direct picosecond laser approach are discussed.