| Previous research has shown, using modified bases in DNA as well as PNA, that there exists a linear relationship between the magnitude of a modified base dipole moment and duplex stability. Past work has been accomplished using 4-substituted phenyl residues in PNA and in DNA; current work focuses on 5-substituted indoles in the context of DNA.; DNA 15-mers were prepared, and each indole residue was then evaluated for its ability to stack within a DNA duplex by conducting thermal denaturation experiments. Thermodynamic parameters Tm, DeltaH, and DeltaS were derived from melting curves using MeltWin software. Using HyperChem modeling software, a number of parameters for the modified nucleosides were calculated. Dipole moment and polarizability were calculated using the PM3 semi-empirical method on energy minimized structures; hydrophobicity and surface area were calculated using the QSAR package. Each of these parameters was plotted against the change in free energy relative to an unsubstituted indole residue. The results indicate a correlation of duplex stability with dipole moment. However, no correlation was observed with any other variables.; The results above were used in the rational design of several universal bases. PNA and DNA monomers of carbazole, 3-nitrocarbazole, and 3,6-dinitrocarbazole were synthesized, incorporated into oligomers, and examined for potential as universal bases. 3,6-Dinitrocarbazole has the highest average Tm of any universal base in PNA; however, its Tm range is larger than expected. When used in the context of DNA, both the mono- and dinitrocarbazoles display superior stability and are non-discriminating. Therefore, applications which require a universal base may benefit from use of these compounds over the traditional bases 5-nitroindole, 3-nitropyrrole, and hypoxanthine.; Two fluorescent bases were also designed and synthesized for use as universal bases in PNA. Both residues were remarkably promiscuous and fairly stable, and displayed an increase in fluorescence intensity when hybridized to complementary DNAs.; Finally, microperoxidase-11 (MP-11) was evaluated for its potential to cause DNA damage in the presence of different reducing agents. Optimal concentration for strand scission was found for both MP-11 and reducing agent, and mechanistic implications are discussed. |
