The effects of sequence context on base dynamics at TpA steps in DNA studied by NMR.

Abstract

Base dynamics, heretofore observed only at TpA steps in DNA, were investigated as a function of sequence context by NMR spectroscopy. The large amplitude conformational dynamics have been previously observed in TnAn segments where n > or = 2. In order to determine whether the dynamic characteristics occur in more general sequence contexts, we examined four self-complementary DNA sequences, [d(CTTTA-NATNTAAAG)2] (where N = A, C, T, G and N = complement of N). The anomalous broadening of the TpA adenine H2 resonance which is indicative of large amplitude base motion was observed in all nine unique four nucleotide contexts. Furthermore, all the adenine H2 resonances experienced a linewidth maximum as a function of temperature, which is a characteristic of the dynamic process. Interestingly, the temperature of the linewidth maximum varied with sequence indicating that the thermodynamics of TpA base dynamics are also sequence dependent. In one example, neither a T preceding nor an A trailing the TpA step was required for base dynamics. These results show that base dynamics, heretofore observed in only a few isolated sequences, occurs at all TpA steps which are either preceded or followed by a thymine or adenine, respectively, and may be characteristic of all TpA steps in DNA notwithstanding sequence context.