TABLE 2.
Specific activities and kinetic parameters of wild-type PEPC and its mutants
| PEPC type/ mutant |
Mean sp acta (U/mg of protein) ± SD | Mean Km (mM) ± SDb |
|
|---|---|---|---|
| KHCO3 | PEP | ||
| Wild type | 47 ± 5 | 2.8 ± 0.18 | 0.62 ± 0.13 |
| R620G | 0.48 ± 0.12 | 3.3 ± 0.22 | 4.88 ± 0.58 |
| K653G | 2.24 ± 0.23 | 3.5 ± 0.28 | 3.31 ± 0.54 |
| K813G | 38 ± 3 | 3.1 ± 0.37 | 0.78 ± 0.21 |
| S869G | 42 ± 3 | 2.9 ± 0.18 | 0.68 ± 0.14 |
| R873G | 43 ± 4 | 2.8 ± 0.25 | 0.66 ± 0.11 |
| N917G | 44 ± 4 | 2.6 ± 0.26 | 0.69 ± 0.22 |
a
All of the enzyme assays were carried out at 25°C. The standard assay contained 100 mM Tris-HCl (pH 8.5), 10 mM KHCO3, 2 mM PEP, 0.1 mM acetyl coenzyme A, 10 mM MgSO4, 0.1 mM NADH, 1.5 IU of malate dehydrogenase, and an appropriate amount of enzyme. All of the assays were repeated three times, and the mean values are shown.
b
The kinetic experiments were carried out under the same condition except for the substrate concentration.