Table 2.
Kinetic parameters of Nudt19 and Nudt7 mutants and chimeras
The experiments were conducted in duplicate as described under “Experimental procedures.” Kinetic constants were determined by nonlinear regression analysis of the data and expressed ± S.E.
| Mutation | Km | kcat |
|---|---|---|
| μm | 1/min | |
| Nudt19 | ||
| WT | 300 ± 40 | 284 ± 12 |
| Nudt19-7-CB | 1134 ± 284 | 0.05 ± 0.01 |
| E112A | N.D.a | N.D. |
| E115A | 1019 ± 226 | 0.16 ± 0.02 |
| E116A | N.D. | N.D. |
| R34M | 680 ± 186 | 0.12 ± 0.01 |
| R39A | 511 ± 64 | 91 ± 4 |
| F40A | 1053 ± 153 | 210 ± 12 |
| L41A | 1250 ± 125 | 101 ± 4 |
| D53A | 434 ± 59 | 323 ± 16 |
| R189A | 1193 ± 184 | 118 ± 11 |
| R189M | 455 ± 80 | 177 ± 10 |
| Nudt7 | ||
| WT | 283 ± 94 | 125 ± 12 |
| Nudt7-19-CB | 1362 ± 328 | 0.28 ± 0.03 |
| Nudt7-19-NB | 836 ± 173 | 0.65 ± 0.04 |
| E92A | N.D. | N.D. |
| E95A | 444 ± 87 | 10 ± 0.7 |
| E96A | N.D. | N.D. |
| R60M | 1010 ± 155 | 0.55 ± 0.03 |
| K65A | 958 ± 277 | 463 ± 56 |
| R66A | 3069 ± 407 | 220 ± 10 |
| R66M | 496 ± 79 | 167 ± 7 |
| D79A | 334 ± 29 | 159 ± 5 |
a N.D. indicates that the parameters were not determined because of undetectable enzymatic activity.