Fig. 2.

Monitoring of real-time DNA relaxation events and step-size distributions. (A) Real-time relaxation events on positively supercoiled DNA are shown at 1.5 pN in the presence of glycerol. An increase in DNA extension upon addition of topo-78 was observed. Each step corresponds to a relaxation event representing one cleavage/religation cycle. The change in linking number is of different size for different events. Hence, topo V relaxes DNA in steps of n supercoils. The dashed lines mark the extension limits used to select events for further analysis in this particular experiment. (B) A step-size distribution of topo-78 acting on a single DNA molecule is shown for events monitored at a force of 0.5 pN in presence of glycerol when 45 plectonemic supercoils were incorporated. A similar shape distribution was observed at all forces studied. The mean step size for each distribution was computed numerically by using the method of Koster et al. (14), yielding a mean ΔLk of 12.3 ± 1.8. (Inset) Step-size distribution for topo-78 measured in absence of glycerol when 45 plectonemic supercoils were applied at a force of 0.5 pN. The line was fit assuming an exponential decay. Error bars represent the square root of the number of events.