Article Figures & Data
Figures
- Figure 1
Effects of endogenous steroids and α-naphthoflavone on CBZ 10,11-epoxidation by expressed CYP3A4 (A) and CYP3A7 (B).
The concentrations of endogenous steroids, α-naphthoflavone, and CBZ were 100 μM. Basal activities of CYP3A4 and CYP3A7 were 3.360 and 0.077 nmol/nmol P450/min, respectively.
- Figure 2
Substrate-velocity curve and Eadie-Hofstee plots of CBZ 10,11-epoxidation by expressed CYP3A4 (A) and CYP3A7 (B).
The reactions were performed in the absence or presence of 100 μM effectors. Lines were drawn using data analyzed by the modified two-site equation for CYP3A4, and by the Michaelis-Menten equation for CYP3A7. ○, CBZ only; ●, plus AND; ▴, plus EST; ▵, plus DHEA-S.
- Figure 3
Substrate-velocity curve and Eadie-Hofstee plots of AND 6β-hydroxylation by expressed CYP3A4.
The reactions were performed in the absence or presence of 100 μM CBZ. Lines were drawn using data analyzed by the modified two-site equation. ○, AND only; ●, plus CBZ.
- Figure 4
Substrate-Velocity curve and Eadie-Hofstee plots of DHEA-S 16α-hydroxylation by expressed CYP3A7.
The reactions were performed in the absence or presence of 100 μM CBZ. Lines were drawn using data analyzed by the Michaelis-Menten equation. ○, DHEA-S only; ●, plus CBZ.
- Figure 5
Kinetic scheme for the effect of endogenous steroids on CYP3A4-mediated CBZ 10,11-epoxidation (A) and the effect of DHEA-S on CYP3A7-mediated CBZ 10,11-epoxidation (B).
C, CBZ; A, AND; E, EST; D, DHEA-S; P, 10,11-epoxi CBZ; P′, 16α-hydroxy DHEA-S.
Tables
- Table 1
Concentration-dependent effects of endogenous steroids on CBZ 10,11-epoxidation by expressed CYP3A4 and CYP3A7
Added Steroids CYP3A4 CYP3A7 μM nmol/nmol P450/min (%) nmol/nmol P450/min (%) none 0 3.275 (100.0) 0.062 (100.0) AND 25 6.229 (190.2) 0.104 (167.6) 50 8.944 (273.1) 0.127 (205.7) 100 9.769 (298.3) 0.134 (216.5) 200 9.320 (284.6) 0.159 (256.6) EST 25 3.249 (99.2) 0.032 (51.8) 50 3.017 (92.1) 0.023 (37.5) 100 2.857 (87.2) 0.023 (36.7) 200 3.085 (94.2) 0.021 (33.8) DHEA-S 25 3.190 (97.4) 0.155 (251.4) 50 2.774 (84.7) 0.184 (296.9) 100 3.163 (108.5) 0.194 (330.9) 200 3.414 (104.3) 0.215 (348.1) The concentration of substrate was 100 μM.
CYP3A4 CYP3A7 Km1 (apparent) Km2(apparent) Vmax2 Vmax2/Km2 Km(apparent) Vmax Vmax/Km μM nmol/nmol P450/min μM nmol/nmol P450/min CBZ only 104.8 248.3 23.64 0.0952 720.9 0.50 0.0007 Plus AND 12.8 139.5 29.03 0.20806 382.8 0.53 0.0014 Plus EST 2.1 666.9 24.76 0.03713 616.7 0.15 0.0002 Plus DHEA-S 80.9 246.4 22.59 0.0917 280.1 0.71 0.0025 Substrate concentrations were 15 to 500 μM. The reaction was performed in the absence or presence of 100 μM effectors.Km1, Km2, andVmax2 were calculated by a modified two-site equation (Vmax1 = 0) V = (Vmax2S2/Km1Km2)/(1 + S/Km1Km2) (Korzekwa et al., 1998)Km and Vmax were calculated from the Michaelis-Menten equation V =Vmax/(1 + (Km/S)).
Km1 (apparent) Km2(apparent) Vmax2 Vmax2/Km2 μM nmol/nmol P450/min AND only 3.5 100.3 117.49 1.17139 Plus CBZ 2.0 97.8 81.37 0.83200 Substrate concentrations were 5 to 500 μM. The reaction was performed in the absence or presence of 100 μM CBZ.
Km(apparent) Vmax Vmax/Km μM nmol/nmol P450/min DHEA-S only 17.0 6.60 0.3882 Plus CBZ 21.5 6.05 0.2814 Substrate concentrations used were 10 to 500 μM. The reaction was performed in the absence or presence of 100 μM CBZ.
