Table 3. Kinetic properties of wild-type and mutant His6–BjHMGS1.
Vmax and Km were measured in the presence of variable concentrations of acetyl-CoA and AcAc-CoA employing a radiometric assay. Hydrolase activity (acetyl-CoA hydrolysis) was measured by converting the remaining [14C]acetyl-CoA to acid stable citrate with excess citrate synthase and oxaloacetate [37]. n.i., no inhibition.
| Enzyme | Vmax (μmol·min−1·mg−1) | Km-acetyl-CoA (μM) | Ki-AcAc-CoA (μM) | Kcat (min−1) (Vmax/[E]) | acetyl-CoA hydrolysis (t1/2) |
|---|---|---|---|---|---|
| Wild-Type | 0.47 | 43 | 38 | 24.9 | 11.1 |
| S89A | 0.14 | 18 | 46 | 7.6 | 18.8 |
| N115A | 0.44 | 27 | 91 | 23.4 | 8.8 |
| R157A | 0.032 | 52 | 242 | 1.7 | 24.1 |
| H188N | 0.056 | 55 | n.i. | 2.9 | 61.7 |
| C212S | 0.078 | 29 | 39 | 4.1 | 14.7 |
| S356A | 0.30 | 26 | 199 | 16.1 | 25 |
| S359A | 4.04 | 24 | 28 | 214.1 | 4.7 |