Table 2. Kinetic parameters for wild-type enzyme and mutant proteins for the full reaction with AIR varieda.
| Vmaxb (μmol min-1mg-1) | Kmb (μM) | Km (fold change) | Kcatc (min-1) | Kcat/Km (μM-1 min-1) | Kcat/Km (fold change) | |
|---|---|---|---|---|---|---|
| wild-type | 55 ± 3 | 22 ± 4 | 0 | 5,000 ± 270 | 230 ± 68 | 1 |
| Y152Ae | 1 ± 0.2 | 460 ± 120 | 18 | 110 ± 20 | 0.50 ± 0.2 | 460 |
| Y152F | 18 ± 0.5 | 68 ± 6 | 3 | 1,600 ± 45 | 24 ± 8 | 8.5 |
| R271A | 0.0013 ± 0.00008 | 254 ± 53 | 12 | 0.11 ± 0.007 | 0.0005 ± 0.0001 | 460,000 |
| R271Q | 0.004 ± 0.0003 | 87 ± 19 | 4 | 0.4 ± 0.03 | 0.004 ± 0.001 | 57,500 |
| R271K | 0.33 ± 0.02 | 608 ± 83 | 28 | 50 ± 2 | 0.076 ± 0.03 | 3,030 |
| H273Ae | 0.41 ± 0.01 | 742 ± 51 | 34 | 26 ± 0.9 | 0.036 ± 0.02 | 6,400 |
| H273Q | 0.0022 ± 0.00008 | 28 ± 4 | 1 | 0.19 ± 0.007 | 0.007 ± 0.002 | 32,900 |
| K353Af | Inactive | nd | nd | nd | nd | nd |
| K353R | Inactived | nd | nd | nd | nd | nd |
a
Reported data are average of duplicates and errors are those obtained from curve fitting (equation 1) the appropriate data set.
b
Steady-state values were determined using the ATP coupled assay system with the concentration of ATP and bicarbonate fixed and the concentration of AIR is varied.
c
Calculated using the molecular weight of the dimer.
d
No measurable activity up to a protein concentration of 700 μg/mL.
e
Kinetic parameters obtained using equation 2.
f
Ref. No (7)