Figure 4.

Calicheamicin γ binds with equal affinity to negatively and positively supercoiled pUC19. The relative affinity of calicheamicin γ for negatively (○) and positively (•) supercoiled pUC19 was determined by a competitive DNA damage assay, as described in Materials and Methods. Calicheamicin γ-induced DNA damage in a [³²P]-labeled DNA fragment was measured in the presence of varying concentrations of unlabeled, supercoiled pUC19. The reduction of damage in the labeled DNA correlates directly with the overall binding constant for calicheamicin with the supercoiled DNA competitor. Representative experimental results are shown in the graph; the average ratio of damage in the presence of negatively supercoiled DNA to that with positively supercoiled DNA is 1.0 ± 0.1 (n = 8).