TABLE 2.
Steady-state kinetic parameters for MJ0499/MJ1277a
| Substrate | Km (μM) | Vmax (U mg−1) | kcat (s−1) | kcat/Km (M−1 s−1) |
|---|---|---|---|---|
| Citraconateb | 80 ± 20 | 15 ± 0.9 | 16 ± 1 | 2.0 × 105 |
| Citraconatec | 40 ± 10 | 13 ± 0.7 | 14 ± 1 | 3.5 × 105 |
| (R)-Citramalateb | 810 ± 80 | 1.5 ± 0.05 | 1.6 ± 0.1 | 2.0 × 103 |
| α-Isopropylmalateb | 1,900 ± 400 | 4.2 ± 0.3 | 4.5 ± 0.3 | 2.4 × 103 |
| β-Isopropylmalateb | 39 ± 7 | 1.8 ± 0.1 | 1.9 ± 0.1 | 5.0 × 104 |
| Maleateb | 400 ± 50 | 34 ± 2 | 36 ± 2 | 9.0 × 104 |
| Maleatec | 180 ± 30 | 19 ± 0.9 | 20 ± 1 | 1.1 × 105 |
a
Kinetic parameters were estimated by nonlinear regression, and the standard errors of those parameters are shown. Turnover numbers were calculated assuming one active site per MJ0499/MJ1277 pair.
b
Initial rates of hydrolyase activity were determined by measuring UV absorbance.
c
Initial rates of hydratase activity were measured in a coupled assay for NADH production using excess IPMDHMj.