The presence of A
n
and A
n
T
n
tracts in double-helical sequences perturbs the structural properties
of DNA molecules, resulting in the formation of an alternate conformation to standard B-DNA known as
B‘-DNA. Evidence for a transition occurring prior to duplex melting in molecules containing A
n
tracts
was previously detected by circular dichroism (CD) and calorimetric studies. This premelting transition
was attributed to a conformational change from B‘- to B-DNA. Structural features of A
n
and A
n
T
n
tracts
revealed by X-ray crystallography include a large degree of propeller twisting of adenine bases, narrowed
minor grooves, and the formation of three-centered H-bonds between dA and dT bases. We report UV
resonance Raman (UVRR) and CD spectroscopic studies of two related DNA dodecamer duplexes,
d(CGCAAATTTGCG)2 (A3T3) and d(CGCATATATGCG)2 [(AT)3]. These studies address the presence
of three-centered H-bonds in the B‘ conformation and gauge the impact of these putative H-bonds on the
structural and thermodynamic properties of the A3T3 duplex. UVRR and CD spectra reveal that the
premelting transition is only observed for the A3T3 duplex, is primarily localized to the dA and dT bases,
and is associated with base stacking interactions. Spectroscopic changes associated with the premelting
transition are not readily detectable for the sugar−phosphate backbone or the cytosine and guanosine
bases. The temperature-dependent concerted frequency shifts of dA exocyclic NH2 and dT C4O vibrational
modes suggest that the A3T3 duplex forms three-centered hydrogen bonds at low temperatures, while the
(AT)3 duplex does not. The enthalpy of this H-bond, estimated from the thermally induced frequency
shift of the dT C4O vibrational mode, is approximately 1.9 kJ/mol or 0.46 kcal/mol.