Figure 5.
Average microscopic parameters of bbTBA blockade. (A) Average second order on rates calculated from kinetic measurements at each voltage as in Fig. 4 were fitted to Eq. 2 either by Igor (dashed line) to yield (parameters ± 95% confidence interval) kON(0) = 1.108 ± 0.3 × 108 M−1s−1 and zδON = 0.012 ± 0.093, or with the final model (solid line) with parameters: kON(0) = 1.0 × 108 M−1s−1 and zδON = 0.05. Data were averaged from six patches; error bars represent the standard deviation of fits of kON vs. [bbTBA] as in Fig. 4 C. (B) Average off rates (±SEM; n = 6 patches) were plotted as a function of voltage and fitted to Eq. 2 (dashed line), yielding parameters (±95% confidence interval) kOFF(0) = 562 ± 27 s−1 and zδOFF = 0.07 ± 0.015. (C) Average microscopic Kd measured from steady-state open probability (filled squares) or kinetics (open squares) were plotted as a function of voltage and fitted with Eq. 2 either by Igor (dashed lines) with parameters (±95% confidence interval): steady-state, Kd = 4.8 ± 1.1 μM and zδ = 0.14 ± 0.08; kinetic, Kd = 5.5 ± 1.1 μM and zδ = 0.11 ± 0.07, or the final model (solid lines) with parameters: steady-state, Kd = 5 μM and zδ = 0.15; kinetic, Kd = 6 μM and zδ = 0.15. (In this particular instance, the Kd in the final model was not constrained to the 8–10 μM range, in order to account for microscopic measurements made earlier in time; see Materials and methods). Error bars represent the standard deviation from fits of the Po vs. [bbTBA] curves as in Fig. 4 D. The Kd for the kinetic measurements was taken to be the intersection where kON = kOFF. Data were averaged from six or four patches for kinetic and steady-state measurements, respectively.