Differential scanning calorimetry (DSC) and infrared spectroscopy (IR) were used to study the thermal behavior of DNA as a function of water content up to 12 wpn (water molecules per nucleotide). The DSC and IR results show cooperativity of thermal DNA denaturation at 12 wpn. In the 3.3 to 7.2 wpn range, an early structural change involving slight disruption of base stacking arises, followed at higher temperature by a denaturation process where the major changes in backbone conformation occur. DNA thermal denaturation in the 3.3 to 12 wpn range is irreversible. However, structural disruptions caused by dehydration are fully reversible. The loss of the DNA ordered conformation upon dehydration leads to a decrease in the endothermic maximum that characterizes the thermal denaturation of DNA. At 3.3 wpn, the DNA loses most of its ordered conformation and undergoes a glasslike transition similar to the transition that denatured DNA experiences during reheating. The calorimetric manifestation of the glass transition is established for denatured DNA at low hydration through the study of water plasticizing effects on T, and enthalpy relaxation. At 1.0 wpn (the lowest water content achieved in this study), the DSC and IR data do not show any thermal or structural transitions, indicating the fully amorphous character of DNA at such low hydration.