TABLE 1.
Kinetic parameters of wild-type and His379 mutant human LVRN/APQs toward various aminoacyl-MCA
Kinetic parameters were determined from Lineweaver-Burk plots. Reactions took place at 37 °C for 5 min.
| Substrate | LVRN/APQ | Kma | kcata | kcat/Km × 103a |
|---|---|---|---|---|
| μm | s−1 | μm−1·s−1 | ||
| Leu-MCA | Wild type | 98.6 ± 19 | 37.7 ± 4.8 | 385 ± 26 |
| H379G | 44.5 ± 15 | 11.3 ± 3.0 | 258 ± 19 | |
| H379F | 119 ± 21 | 49.8 ± 6.4 | 419 ± 24 | |
| H379K | 109 ± 9.6 | 33.7 ± 2.7 | 308 ± 3.2 | |
| H379L | 28.1 ± 2.7 | 4.85 ± 0.22 | 173 ± 11 | |
| Met-MCA | Wild type | 65.0 ± 5.0 | 10.9 ± 1.0 | 168 ± 4.3 |
| H379G | 26.4 ± 6.3 | 3.59 ± 0.66 | 137 ± 10 | |
| H379F | 62.8 ± 19 | 18.1 ± 3.9 | 292 ± 22 | |
| H379K | 61.5 ± 1.9 | 11.6 ± 2.7 | 189 ± 5.5 | |
| H379L | 9.26 ± 0.93 | 1.70 ± 0.066 | 184 ± 11 | |
| Arg-MCA | Wild type | 21.0 ± 5.1 | 4.48 ± 0.49 | 219 ± 33 |
| H379G | 11.1 ± 1.5 | 3.11 ± 0.23 | 282 ± 20 | |
| H379F | 21.5 ± 1.1 | 5.74 ± 0.17 | 267 ± 5.9 | |
| H379K | 18.9 ± 2.4 | 1.62 ± 0.12 | 84.9 ± 4.5 | |
| H379L | 9.85 ± 1.2 | 2.07 ± 0.17 | 210 ± 8.5 | |
| Lys-MCA | Wild type | 31.2 ± 1.6 | 4.83 ± 0.19 | 155 ± 2.0 |
| H379G | 15.5 ± 5.4 | 4.10 ± 1.2 | 269 ± 22 | |
| H379F | 31.3 ± 8.0 | 6.68 ± 1.5 | 215 ± 9.5 | |
| H379K | 27.5 ± 1.2 | 4.31 ± 0.14 | 157 ± 2.7 | |
| H379L | 5.82 ± 0.23 | 1.82 ± 0.044 | 313 ± 7.0 | |
| Phe-MCA | Wild type | 43.2 ± 4.5 | 2.19 ± 0.18 | 50.9 ± 1.1 |
| H379G | 26.1 ± 2.7 | 0.862 ± 0.062 | 33.2 ± 1.2 | |
| H379F | 44.7 ± 7.4 | 3.17 ± 0.56 | 71.0 ± 5.5 | |
| H379K | 44.2 ± 2.4 | 1.03 ± 0.11 | 23.1 ± 1.5 | |
| H379L | 18.2 ± 4.4 | 1.74 ± 0.22 | 97.7 ± 10 |
a Values are the mean ± S.D. (n = 3).