TABLE 1.
LTB4, AA and Oleate ω-Hydroxylation by Recombinant Human CYP4F2 and Liver Microsomes
| SUBSTRATE | |||
|---|---|---|---|
| Enzyme | LTB4a | AAb | Oleateb |
| CYP4F2 | 0.20 | 2.6 ± 0.9 | 5.1 |
| CYP4F2 Supersomes | < 0.1 | 3.8 | 21.6 |
| CYP4F3b | 0.22 | 2.4 | 4.7 |
| CYP4F3b Supersomes | 2.13 ± 1.6 | 16.9 | 33.0 |
| CYP4F12 Prep #1 | nd | < 0.1 | < 0.1 |
| CYP4F12 Supersomes | < 0.1 | < 0.1 | < 0.1 |
| Liver Microsomes Subject A | 0.56 | 2.4 ± 0.2 | 2.1 |
| Liver Microsomes Subject B | 0.73 ± 0.1 | 6.9 | 2.0 ± 0.9 |
LTB4, AA and oleate ω-hydroxylation were assessed in incubation mixtures (0.25 ml) containing: a) reconstituted systems [25–50 pmol CYP4F enzyme, 75–150 pmol P450 reductase, 7.5–15 µg DLPC and 100–200 pmol b5; b) supersomes® from T. ni insect cells expressing human CYP4F2, CYP4F3b or CYP4F11 (protein equivalent to 25 pmol P450) or; c) human liver microsomes (protein equivalent to 50–150 pmol P450). Other incubation components included 100 mM potassium phosphate buffer (pH 7.4), 0.5 mM NADPH, and LTB4 (30 µM), AA (100 µM) or oleate (100 µM). Reactions were initiated with NADPH, and were terminated after 10 – 15 min at 37°C. Formati on of 20-OH LTB4, 20-HETE and 18-OH oleate were then assessed by HPLC as described in Materials and Methods.
Data expressed as pmol product formed/min/nmol P450, and denote the average of at least 3 individual determinations. Results given as the mean ± SD represent 9–24 individual determinations.
Data expressed as nmol product formed/min/nmol P450, and denote the average of at least 3 individual determinations. Results given as the mean ± SD represent 6–15 individual determinations.