Kinetic parameters for 4-MU O-glucuronidation by UGT1A9, UGT1A7, UGT1A8, and UGT1A10 expressed in insect cells (Supersomes) and wild-type and mutants UGT1A9 expressed in HEK293 cells
The substrate concentrations were 0.5 to 100 μM for HEK293 expression systems and UGT1A7 and UGT1A9 Supersomes and 25 to 1000 μM for UGT1A8 and UGT1A10 Supersomes. Data are the mean ± S.D. of three independent determinations. The Vmax values were normalized with the relative UGT expression level.
Km | Vmax | Vmax/Km | |
|---|---|---|---|
| μM | nmol/min/unit | μl/min/unit | |
| Supersomes | |||
| UGT1A9 | 9.6 ± 1.3 | 1.2 ± 0.1 | 123 ± 14 |
| UGT1A7 | 23.9 ± 3.9 | 1.4 ± 0.3 | 57 ± 3 |
| UGT1A8 | 191.7 ± 8.8 | 0.2 ± 0.01 | 1.1 ± 0.1 |
| UGT1A10 | 76.2 ± 14.6 | 0.02 ± 0.002 | 0.28 ± 0.05 |
| HEK293 cells | |||
| Wild-type UGT1A9 | 4.4 ± 0.5 | 3.9 ± 0.3 | 891 ± 45 |
| Mut 1 | 10.1 ± 0.3** | 7.3 ± 1.1** | 723 ± 109 |
| Mut 2 | 3.6 ± 0.5 | 3.2 ± 0.3 | 913 ± 176 |
| Mut 3 | 5.6 ± 0.6 | 10.4 ± 2.1** | 1883 ± 427** |
| Mut 4 | 9.1 ± 2.2** | 12.8 ± 1.0** | 1441 ± 242 |
| Mut 5 | 2.1 ± 0.4 | 5.5 ± 0.2 | 2678 ± 456** |
| Mut 6 | 5.9 ± 1.3 | 4.0 ± 0.4 | 681 ± 78 |
| Mut 7 | 4.4 ± 1.1 | 3.4 ± 0.6 | 789 ± 73 |
↵** P < 0.01 compared with wild-type UGT1A9.