Ultraviolet absorbance methods were used to characterize the thermodynamics of melting of a series of 16 bp deoxyoligonucleotides over a wide range of NaCl concentrations (0-4.5 M) and to obtain complete thermodynamic profiles for their melting at 0.115 and 4.5 M NaCl. The sequence of the series (one strand of duplex) was: 5'-CGCGCGCGAMNGACTG-3', where C indicates m5dC and -MN- was varied to include all combinations of Py:Py stacks (CC, TT, CT, TC). The unmethylated deoxyoligonucleotide 5'-CGCGCGCGACTGACTG-3' was used as a control sequence. All of the methylated oligonucleotides studied undergo a NaCl-induced transition to a hybrid form containing a left-handed, Z-DNA, region joined to a right-handed region by a B-Z junction. Our experiments allowed us to quantitatively evaluate the effects of NaCl, sequence, and methylation and the transition to the hybrid BZ structure on DNA thermal stability. We found that alteration of a single dinucleotide step has profound effects on the thermal stabilities of the 16 bp fragments studied. Methylation was found to destabilize the double helix, resulting in a decrease in Tm. Transition to the hybrid BZ structure, somewhat surprisingly, was found to only slightly destabilize DNA, with an observed decrease in free energy of melting of approximately 0.5 kcal/mol relative to the control, right-handed, sequence in high salt. Transition melting temperatures (Tm) were found, in agreement with previous studies on polymeric DNA, to depend upon NaCl concentration in a complicated, nonlinear fashion. m values increase to maximal values at circa 1.0 M NaCl, but decrease thereafter with further addition of salt.(ABSTRACT TRUNCATED AT 250 WORDS)