Table 2.
Comparison of apparent kinetic parameters for reaction of the WT and mutants of AtHPPD at pH 7.0∗. Each experiment was carried out in triplicate.
| Enzyme | k cat (s−1) | K m (μM) | k cat/Km |
|---|---|---|---|
| WT | 1.079 ± 0.053 | 1.87 ± 0.37 | 0.5770 ± 0.1142 |
| N423A | 0.545 ± 0.061 | 15.66 ± 4.29 | 0.0348 ± 0.0095 |
| F381A | 0.924 ± 0.032 | 9.10 ± 0.94 | 0.1015 ± 0.0105 |
| Q293A | 0.014 ± 0.001 | 90.41 ± 11.33 | 0.00015 ± 0.00001 |
| Q307A | 0.176 ± 0.004 | 30.31 ± 1.67 | 0.0058 ± 0.0003 |
| S267A | 0.249 ± 0.025 | 5.33 ± 1.59 | 0.0467 ± 0.0139 |
| N282A | 0.039 ± 0.003 | 10.83 ± 2.33 | 0.0036 ± 0.0008 |
∗ Reactions were conducted in air-saturated 20 mM HEPES (pH 7.0) at 25°C and kinetically analyzed as described in Experimental Procedures. Errors for individual parameters were obtained from the nonlinear regression fit of the data to the Michaelis-Menten equation.