Table 1.
Anaerobic Characterization of H49 and H299 Variantsa
| enzyme | Kmapp,d-GAP (μM) | Kmapp,pyruvate (μM) | kcat (min−1) | kcat/Kmd-GAP (min−1 μM−1) | kcat/Kmpyruvate (min−1 μM−1) |
|---|---|---|---|---|---|
| wild type | (1.42 ± 0.04) × 10 | (4.42 ± 0.07) × 10 | (8.29 ± 0.2) × 10 | (5.7 ± 0.2) × 100 | (1.93 ± 0.09) × 100 |
| H49A | (1.62 ± 0.07) × 10 | (1.07 ± 0.02) × 103 | (2.69 ± 0.05) × 10 | (1.64 ± 0.05) × 100 | (2.53 ± 0.01) × 10−2 |
| H49N | (2.9 ± 0.1) × 10 | (1.61 ± 0.08) × 103 | (3.7 ± 0.2) × 10 | (1.13 ± 0.02) × 100 | (2.5 ± 0.1) × 10−2 |
| H299Ab | (3.30 ± 0.4) × 100 | (1.10 ± 0.01) × 103 | (2.7 ± 0.4) × 100 | (9.0 ± 1) × 10−1 | (2.22 ± 0.04) × 10−3 |
| H299N | (1.50 ± 0.04) × 10 | (1.08 ± 0.08) × 103 | (1.04 ± 0.04) × 10 | (6.5 ± 0.2) × 10−1 | (1.04 ± 0.06) × 10−2 |
a
The error represents the standard error, where n = 3 (Km and kcat/Km) or n = 6 (kcat).
b
Michaelis–Menten conditions not achievable due to the high DXPS concentration (2 μM) required to observe activity.