Point mutations change the thermal denaturation profile of a short DNA fragment containing the lactose control...

To understand the denaturation process of short DNA segments we have chosen a 203-base pair (bp) restriction fragment containing the lactose control region. A steady decrease in GC content exists between its i proximal and z proximal ends. We confirm that this fragment melts at low salt in two subtransitions. A GC to AT mutation in the AT-rich region (mutation UV5) increases the number of denatured base pairs in the first subtransition and decreases the cooperativity of the melting process. A GC to AT mutation in the GC-rich region (mutation L8) decreases the number of denatured base pairs in the first subtransition and increases the cooperativity. These mutations induce the same shift in the temperature of half denaturation. The effects of both mutations are additive. A short deletion at the z end of the fragment affects only the first subtransition. When four GC pairs are added to both end, the fragment melts in one transition. Comparison with the results obtained with a larger 789-bp lac fragment reveals strong end effects on base pair stability and suggests that denaturation of the 203-bp fragment proceeds unidirectionally from the z end. Good agreement is shown with the predictions made with the "z ipper model" of Crothers et al. (1965).