Figure 7.

dmant-ADP binding to p-huM3AMD and acto-P-huM3AMD. (A) Rate of binding of dmant-ADP to p-huM3AMD as a function of nucleotide concentration. The observed rates (k_obs) were obtained by fitting the fluorescence transients to double exponentials. The apparent second-order rate constants for binding of dmant-ADP to p-huM3AMD are 0.87 μM⁻¹ s⁻¹ for the fast phase (○) and 0.002 μM⁻¹ s⁻¹ for the slow phase (●). The dissociation rates of dmant-ADP determined from the y-intercept of the fast phase and slow phase are 8.6 and 0.12 s⁻¹, respectively. The inset shows a typical recording of the binding of dmant-ADP (5 μM) to 0.5 μM p-huM3AMD. The solid line is the best fit to double-exponential kinetics with a fast k_obs of 11.13 s⁻¹ and a slow k_obs of 0.13 s⁻¹. (B) Rate of binding of dmant-ADP to acto-p-huM3AMD as a function of nucleotide concentration. The observed rates (k_obs) were obtained by fitting the fluorescence transients to a single exponential. The apparent second-order rate constant for binding of dmant-ADP to acto-p-huM3AMDis 0.56 μM⁻¹ s⁻¹. The dissociation rate of dmant-ADP determined from the y-intercept is 8.1 s⁻¹. The error bars represent the standard error from three independent experiments. For several data points, the error bars are within the circles. The inset shows a typical recording of the binding of dmant-ADP (5 μM) to 0.5 μM acto-p-huM3AMD. The solid line is the best fit to single-exponential kinetics with a k_obs of 10.41 s⁻¹.