Table 1.
Determination of binary complex parameters using either CPR-K56Q or P450 2E1 as the titrant.
| Method of analysis | CPR-K56Q as the titrant | P450 2E1 as the titrant | ||
|---|---|---|---|---|
| Kd (nM) | kcat (min−1) | Kd (nM) | kcat (min−1) | |
| Independenta | 30 ± 10 | 9.7 ± 1.2 | 12 ± 5 | 7.9 ± 0.2 |
| Globalb | 36 ± 3 | 10.7 ± 0.3 | 15.6 ± 1.4 | 7.2 ± 0.2 |
a
Kd values were determined by averaging the results from the fit of each titration curve to the respective binding quadratic equation (Equation 1). The turnover numbers, kcat, were derived from slope of the linear regression between Vmax and the concentration of limiting complex partner as shown in Figure 1 and Figure 2, Panel B.
b
Parameters reflect the global fit of all three titration curves to a binary complex mechanism using the software DynaFit (23). The input data for this analysis is included in Supplemental Information.