Recognition and Visualization of Nucleic Acids
Doctoral thesis, 2003
The work presented in this Thesis focuses on the interaction between nucleic acids and small molecules, as well as binding or incorporation of reporter chromophores into nucleic acids. The results could help in studying and understanding processes that are fundamental to describe mechanisms behind a genetic disorder. For example, the molecules investigated could be used for studying the mechanism of interaction between DNA and other molecules, such as proteins or drugs, and also give information about intrinsic properties of DNA during, for example, DNA-protein interaction.
In Paper I, a dramatic color change of a cyanine dye, 3,3'-diethylthiadicarbocyanine, is used in the development of a fast and straightforward low-cost assay for genetic screening. The aggregation of several dyes within the minor groove of the double helix, responsible for the color change of the dye upon binding to PNA-DNA duplexes, is also in itself a very interesting observation, giving information about the molecular dimensions of the binding site.
Paper II and III reports on thermodynamic and kinetic DNA-binding properties of the dimeric compound, [μ(11,11'-bidppz)(phen)4Ru2]4+. Flow linear dichroism and luminescence studies show that the ruthenium complex extremely slowly rearranges from an initial groove-bound to a final threaded intercalated geometry. The slowness of this rearrangement of DNA-binding modes, due to the requirement of opening of at least on base pair, is to the best of our knowledge, unprecedented. In Paper IV, the SDS-sequestering method for measuring dissociation of cationic drugs from DNA is reported to strongly overestimate the rate of dissociation of bulky multicationic intercalators. An explanation, where the surfactant molecules are involved in a reduction of the activation barrier for the cationic intercalators to leave DNA, is presented as well as an alternative, more reliable method for determining dissociation rates based on use of dummy DNA as an absorbent for the dissociated species.
In Paper V and VI, a novel fluorescent base analogue, tC, is presented. Its high fluorescence quantum yield (0.2) is preserved upon incorporation into nucleic acid and subsequent hybridization to a complementary DNA strand. The minimal influence of environment on the photophysical properties of tC and its firm stacking between the bases, makes it very promising as a probe of intra- and inter-molecular distance estimations in nucleic acid systems.
fluorescent base analogue