TABLE 1.
Kinetic parameters
| Parameter | Notes | Fujioka et al. | This study |
|---|---|---|---|
| Mektot | Total cellular Mek concentration | 1.4 μM | 1 μM |
| Erktot | Total cellular Erk concentration | 0.96 μM | 10 μM |
| kon,Raf-Mek | Association rate constant of Raf-Mek complex | 0.65 μM−1 s−1 | 0.65 μM−1 s−1 |
| koff,Raf-Mek | Dissociation rate constant of Raf-Mek complex | 0.065 s−1 | 0.065 s−1 |
| kcat,Raf-Mek | Catalytic turnover constant of Raf-Mek complex | 0.18 s−1 | 0.18 s−1 |
| (Vmax/KM)Mek-PPase | First-order rate constant of Mek-phosphatase | 0.01 s−1 | (0.01 s−1) |
| Vmax,Mek-PPase | Maximal velocity of Mek-phosphatase | — | 0.001 μM s−1 |
| KM,Mek-PPase | Michaelis-Menten constant of Mek-phosphatase | — | 0.1 μM |
| kon,Mek-Erk | Association rate constant of Mek-Erk complex | 0.88 μM−1 s−1 | 0.88 μM−1 s−1 |
| koff,Mek-Erk | Dissociation rate constant of Mek-Erk complex | 0.088 s−1 | 0.088 s−1 |
| kcat,Mek-Erk | Catalytic turnover constant of Mek-Erk complex | 0.22 s−1 | 0.22 s−1 |
| (Vmax/KM)Erk-PPase | First-order rate constant of Erk-phosphatase | 0.014 s−1 | (0.08 s−1) |
| Vmax,Erk-PPase | Maximal velocity of Erk-phosphatase | — | 0.04 μM s−1 |
| KM,Erk-PPase | Michaelis-Menten constant of Erk-phosphatase | — | 0.5 μM |
The total protein concentrations and kinetic parameters of the model depicted in Fig. 1 D are listed under the heading “This study”, and compared to the values measured by Fujioka et al. (26). Fujioka et al. estimated the apparent first-order rate constant (Vmax/KM) of Mek and Erk dephosphorylation only. We assumed saturated Michaelis-Menten kinetics in the model, because 1), the time course data in Fujioka et al. (26) indicates saturation in the dephosphorylation reactions; and 2), the KM values of phosphatases toward full-length substrates are frequently in the submicromolar range (52,53). See Supplementary Material for differential equations of the model.