SynopsisReversible and irreversible conformational changes in the acid-induced denaturation of DNA were studied by spectrophotometric titration, sedimentation, and melting measurements. A GC-rich DNA (72 mole-%) shows complete or partial reversibility of the titration profiles within the pH region of transition from helix to coil, while AT-rich DNA (29 mole-%) is irreversible in its titration behavior a t each acid pH below the onset of the transition. The results for GC-rich DNA further indicate distinct differences in the titration behavior, which can be attributed to differences in the frequency of GC clusters along the UTA molecule. Plots of the sedimentation coefficient arid the parameter ua;p against pH lead to the conclusion that conformational changes occur before the onset of the acid-induced helix-coil transition. These alterations are more pronounced upon protonation of larger GC-rich domains than of smaller ones, as concluded from very marked differences observed in the sedimentation-pH behavior of two GC-rich DXA's. An acid denaturation scheme for a GC-rich DNA segment is suggested. Reversihility of the acid denaturation is explained by the existence of stable, protonated, single GC base pairs in nonprotonated stacked singlestranded domains formed in the acid-induced transition region.