Table 1.
Steady-State Kinetic Parameters for Wild-Type and Y254F Flavocytochromes b2
| kinetic parametera | wild-type enzymeb | Y254F |
|---|---|---|
| Vmax, s−1 | 372 ± 12 | 13.2 ± 0.4 |
| V/Klactate, mM−1 s−1 | 2300 ± 215 | 44 ± 5.0 |
| Klactate, mM | 0.16 ± 0.02 | 0.28 ± 0.02 |
| DV | 2.9 ± 0.1 | 5.0 ± 0.3c |
| D(V/K) | 3.0 ± 0.3 | 5.0 ± 0.3 |
| D2OV | 1.38 ± 0.07 | 1.54 ± 0.07 |
| D2O(V/K) | 0.90 ± 0.04 | 1.54 ± 0.07 |
| DVD2O | ndd | 4.5 ± 0.3 |
| D(V/K)D2O | nd | 4.5 ± 0.3 |
| D2OVD | 1.1 ± 0.03 | 1.41 ± 0.05 |
| D2O(V/K)D | 0.87 ± 0.1 | 1.41 ± 0.05 |
a
Conditions: 100 mM potassium phosphate, 1 mM EDTA, and 1 mM ferricyanide, pH 7.5 or pD 8.0, 25 °C.
b
From ref 21.
c
For all of the isotope effects for the Y254F enzyme, the data fit better to eq 2, which describes identical isotope effects on the V and V/K values, than to eq 1, which yields different DV and D(V/K) values.
d
Not determined.