TABLE 3.
Kinetic parameters of purified wild-type and mutant subtilisins for hydrolysis of AAPF at 10°Ca
| Sample | kcat (s−1) | Km (μM) | kcat/Km (105 s−1 M−1) | Value relative to wild type |
|---|---|---|---|---|
| Wild type | 29.3 (±1) | 96.1 (±3) | 3.0 | 1.0 |
| G131D | 33.1 (±3) | 76.5 (±5) | 4.4 | 1.4 |
| G131W | 34.5 (±2) | 75.4 (±0) | 4.5 | 1.5 |
| G131M | 27.5 (±3) | 48.4 (±1) | 5.7 | 1.8 |
| G131R | 31.4 (±2) | 53.7 (±0) | 5.9 | 1.9 |
| G131F | 41.9 (±2) | 54.6 (±2) | 7.6 | 2.5 |
a
Enzyme activity was assayed using acetone-precipitated sample by the method of Wells et al. (29) with a slight modification. AAPF (Sigma) was used as the substrate. Estimation of enzyme concentration was carried out by active-site titration with SSI.