Kinetic analysis of glucuronide formation for BaP metabolites for UGT1A10139Glu- and UGT1A10139Lys-overexpressing cell homogenates All the reactions were performed using 0.5 mg of UGT1A10-overexpressing cell homogenate, with incubations performed for 1 h. Kinetic data are reported as mean ± S.D. for three independent experiments. Km represents apparent Km.
Substrate | UGT1A10139Glu | UGT1A10139Lys | ||||||||
|---|---|---|---|---|---|---|---|---|---|---|
| Km | Vmax
a
| Vmax/Kma | Km | Vmax
a
| Vmax/Kma | |||||
| μM | nmol · min–1 · mg–1 | μl · min–1 · mg–1 | μM | nmol · min–1 · mg–1 | μl · min–1 · mg–1 | |||||
| BPD | 46.8 ± 4.2 | 7.7 ± 0.4 | 165 ± 6.5 | 53.2 ± 11 | 3.9 ± 0.2b | 73.3 ± 13b | ||||
| 3-OH-BaP | 9.7 ± 1.2 | 24.8 ± 1.9 | 2557 ± 124 | 15.4 ± 1.6b | 17.7 ± 1.2b | 1149 ± 41b | ||||
| 7-OH-BaP | 9.8 ± 0.5 | 25.8 ± 0.7 | 2633 ± 81 | 9.6 ± 0.5 | 13.2 ± 0.8b | 1375 ± 17b | ||||
| 9-OH-BaP | 38.2 ± 6.2 | 69.4 ± 9.2 | 1817 ± 59 | 48.2 ± 3.6 | 44.8 ± 2.4 | 929 ± 21b | ||||
| 1-OH-pyrene | 11.3 ± 3.3 | 90.0 ± 2.4 | 7965 ± 2100 | 14.7 ± 3.9c | 50.4 ± 3.1b | 3429 ± 770c | ||||
↵ a Vmax values are adjusted per milligram of the corresponding UGT1A protein as determined by Western blot analysis
↵ b Significant (p < 0.01b or p < 0.05c) increase in kinetic parameter observed for homogenates from UGT1A10139Glu- versus UGT1A10139Lys-overexpressing cells
↵ c Significant (p < 0.01b or p < 0.05c) increase in kinetic parameter observed for homogenates from UGT1A10139Glu- versus UGT1A10139Lys-overexpressing cells