Table 3. Enzyme kinetic parameters of UGTs 2A1, 2B4, 2B7, 2B15, and 2B17-catalyzed glucuronidation in the absence and presence of BSA.
| No BSA | 0.1% BSA | |||||
| UGT enzyme/substrate | K m or S 50± S.E. (95% CI) | V max ± S.E. (95% CI) | Model, r 2 | K m or S 50± S.E. (95% CI) | V max ± S.E. (95% CI) | Model, r 2 |
| µM | nmol•min–1•mg–1 | µM (S.E.) | µM | nmol•min–1•mg–1 | µM (S.E.) | |
| UGT2A1 | ||||||
| 6-Hydroxyindole | 1474±121(1223–1724) | 0.79±0.04(0.72–0.87) | MM, 0.99 | 56.6±2.3***(51.8–61.4) | 1.2±0.01***(1.17–1.22) | MM, 0.99 |
| 4-MU | 48.1±4.8(38.2–57.9) | 2.1±0.1(1.9–2.1) | MM, 0.97 | 2.73±0.20†(2.41–3.05) | 2.1±0.02†(2.1–2.2) | SI, 0.97;K i = 2045 (255) |
| UGT2B4 | ||||||
| 17α-Estradiol (17-glucuronide) | 4.51±0.68(3.09–5.93) | 0.053±0.002(0.048–0.058) | MM, 0.95 | 1.60±0.23***(1.12–2.08) | 0.047±0.002(0.044–0.050) | MM, 0.91 |
| 1-Naphthol | 210±12(186–235) | 0.025±0.001(0.023–0.027) | HE, 0.96;h = 2.62 (0.36) | 99.8±7.5***(84.3–115.4) | 0.0080±0.0032***(0.0074–0.0087) | HE, 0.89;h = 4.13 (1.00) |
| 4-MU | 298±43(210–387) | 0.043±0.002(0.039–0.047) | MM, 0.95 | 192±15*(161–222) | 0.035±0.001***(0.033–0.037) | MM, 0.98 |
| UGT2B7 | ||||||
| 17α-Estradiol (17-glucuronide) | 0.128±0.015(0.098–0.158) | 0.014±0.0003(0.014–0.015) | MM, 0.96 | 0.0379±0.0065***(0.0242–0.0515) | 0.016±0.0003***(0.016–0.017) | MM, 0.97 |
| 17β-Estradiol (17-glucuronide) | 2.56±0.29(1.96–3.16) | 0.0067±0.0003(0.0062–0.0072) | MM, 0.96 | 0.832±0.096***(0.633–1.031) | 0.0063±0.0002(0.0059–0.0067) | MM, 0.94 |
| 4-MU | 301±65(165–436) | 0.28±0.03(0.22–0.34) | HE, 0.93; h = 1.75(0.46) | 212±17(177–247) | 0.39±0.01*(0.36–0.42) | HE, 0.99;h = 2.01 (0.24) |
| UGT2B15 | ||||||
| 17α-Estradiol (3-glucuronide) | 10.2±1.0(8.0–12.3) | 0.069±0.003(0.062–0.076) | SI, 0.99;K i = 360 (92) | 5.58±0.56***(4.24–6.74) | 0.085±0.003**(0.077–0.092) | SI, 0.98;K i = 348 (80) |
| 4-MU | 29.6±0.9(27.6–31.5) | 0.83±0.01(0.82–0.85) | MM, 0.99 | 7.06±0.30***(6.54–7.54) | 0.53±0.01***(0.53–0.54) | MM, 0.99 |
| UGT2B17 | ||||||
| 17β-Estradiol (17-glucuronide) | 6.01±0.26(5.47–6.54) | 0.013±0.0002(0.0125–0.0132) | MM, 0.99 | 4.01±0.24***(3.51–4.51) | 0.013±0.0002(0.0126–0.0134) | MM, 0.98 |
| 1-Naphthol | 540±26(485–595) | 0.28±0.01(0.26–0.31) | MM, 0.99 | 484±27(427–541) | 0.34±0.02**(0.30–0.37) | MM, 0.99 |
| 4-MU | 377±42(291–463) | 0.050±0.002(0.049–0.058) | MM, 0.99 | 383±29(322–444) | 0.068±0.002 ***(0.064–0.072) | MM, 0.99 |
The glucuronidation rates are presented as expression level-normalized values ± S.E., except for the commercial UGT2B15. The 95% CI are presented in the parenthesis. The p-values were calculated using the extra sum-of-squares F-test (see Materials and Methods for all details).
MM, Michaelis-Menten; SI, substrate inhibition; HE, Hill equation;
*
P<0.05;
**
P<0.01;
***
P<0.001.