Binding of ethidium derivatives to natural DNA: a 300 MHz 1H NMR study

The binding of three ethidium derivatives, ethidium (1), des-3-amino ethidium (2) and des-8-amino ethidium (3), to short (approximately 35 base pairs), random sequence DNA has been investigated using 300 MHz proton NMR. At 35 degrees C all three drugs cause upfield shifts of the resonances from the exchangeable imino protons, as expected for intercalative binding to DNA. However, the lineshapes vary significantly with the nature of the drug. The temperature dependence of the spectra of the DNA shows that differences between spectra observed at 35 degrees C with ethidium and with des-3-amino ethidium are primarily due to differences in the drug binding kinetics rather than to differences in mode of binding. Removal of the amino group at position 3, but not at position 8, on the parent ethidium shortens the lifetime of the intercalative state; this implies that the 3-NH2 group is involved in stabilization of the drug-DNA complex. Analysis of the drug-DNA spectra indicates that there is a preference for binding of the drugs adjacent to G.C base pairs.