Figure 6.
Direct measurement of εA-DNA dissociation from Y127W and Y159W AAG. Double-mixing experiments were performed using stopped-flow fluorescence. The AAG•DNA complex was formed, aged for one second, and then chased with an excess of pyrrolidine-DNA as a competitor. The normalized fluorescence was obtained by dividing the observed fluorescence signal by the fluorescence of the sample after the dissociation reaction was complete. The traces reflect the average of 3 separate mixing experiments and the solid lines depict the best fit of a single exponential. Only one out of every twenty data points is shown for clarity. The observed rate constants for dissociation are 0.12 s-1 for Y127W (◇) and 0.37 s-1 for Y159W (○) mutant proteins.