TABLE 1.
Apparent kinetic parameters for SalR and its mutated versions
| Version | Km | Kim | Vmax/opt | kcat | kcat/Km | Km NADPH |
|---|---|---|---|---|---|---|
| μm | μm | nkat mg−1 | s−1 | s−1mm−1 | μm | |
| SalR-WT | 2.1 ± 0.6 | 184 ± 47 | 35.61 ± 2.82a | 1.306 ± 0.104 | 661.7 | 3.5 ± 0.8b |
| F104A | 31.8 ± 3.6 | 466 ± 64 | 74.38 ± 4.91a | 2.727 ± 0.180 | 86.2 | 9.7 ± 2.5c |
| V106A | 8.9 ± 2.9 | 283 ± 90 | 67.13 ± 9.03a | 2.458 ± 0.331 | 295.4 | 7.0 ± 2.3c |
| D107A | 14.8 ± 2.4 | 436 ± 78 | 16.61 ± 1.11a | 0.610 ± 0.040 | 41.7 | 30.5 ± 6.0c |
| S181A | 5.0 ± 1.9 | 199 ± 60 | 52.75 ± 6.62a | 1.933 ± 0.243 | 430.3 | 4.5 ± 1.7d |
| T182A | 4.1 ± 1.2 | 58 ± 15 | 3.43 ± 0.39a | 0.126 ± 0.014 | 32.4 | 21.3 ± 5.1d |
| L185A | 13.2 ± 1.7 | NDe | 0.44 ± 0.01f | 0.016 ± 0.001 | 1.2 | 29.7 ± 6.7g |
| Ser | 1.1 ± 0.2 | 377 ± 76 | 4.03 ± 0.15a | 0.148 ± 0.006 | 137.9 | 12.9 ± 4.1d |
| Val | 2.1 ± 0.6 | 135 ± 30 | 18.69 ± 1.34a | 0.685 ± 0.049 | 347.6 | 14.2 ± 3.8d |
| K186V | 7.8 ± 2.1 | 18 ± 8 | 27.23 ± 8.05a | 0.998 ± 0.295 | 149.3 | 3.4 ± 1.1d |
| L266A | 49.4 ± 2.2 | ND | 13.68 ± 0.17f | 0.501 ± 0.006 | 10.2 | 15.5 ± 1.3g |
| M271A | 22.5 ± 2.8 | ND | 0.16 ± 0.01f | 0.006 ± 0.001 | 0.3 | 19.8 ± 5.4g |
| N272A | 90.1 ± 5.0 | ND | 0.57 ± 0.01f | 0.021 ± 0.001 | 0.2 | 15.6 ± 3.4g |
| I275A | 33.1 ± 5.9 | 1249 ± 330 | 87.03 ± 7.58a | 3.188 ± 0.278 | 98.0 | 21.1 ± 1.9c |
| Val | 2.1 ± 0.9 | 122 ± 41 | 27.89 ± 3.01a | 1.022 ± 0.110 | 568.5 | 4.7 ± 0.7c |
| F104A/I275A | 408.6 ± 6.6 | ND | 66.85 ± 0.49f | 2.450 ± 0.017 | 5.9 | 43.0 ± 12.5h |
a Vopt.
b All kinetic parameters for salutaridine were recorded at 250 μm NADPH; salutaridine concentrations for the determination of NADPH affinity were 10 μm.
c All kinetic parameters for salutaridine were recorded at 250 μm NADPH; salutaridine concentrations for the determination of NADPH affinity were 75 μm.
d All kinetic parameters for salutaridine were recorded at 250 μm NADPH; salutaridine concentrations for the determination of NADPH affinity were 15 μm.
e ND, not detectable up to a salutaridine concentration of 1000 μm.
f Vmax.
g All kinetic parameters for salutaridine were recorded at 250 μm NADPH; salutaridine concentrations for the determination of NADPH affinity were 500 μm.
h All kinetic parameters for salutaridine were recorded at 250 μm NADPH; salutaridine concentrations for the determination of NADPH affinity were 1000 μm.