Inhibition of purified and membrane-bound flavin-containing monooxygenase 1 by (N,N-dimethylamino)stilbene carboxylates.
(E)-[2-(4-(Dimethylamino)phenyl)vinyl]benzenes bearing a nitrile or carboxyl group in the 2', 3', or 4' position were synthesized and tested for substrate activity with purified pig liver flavin-containing monooxygenase (FMO1). Although the nitrile derivatives were too insoluble to saturate the catalytic site at pH 7.4, they appeared to be substrates with K(m)'s somewhat above their maximum solubility (approximately equal to 0.1 mM) in the assay medium. Of the three carboxylic acid analogs, (E)-4-[2-(4(dimethylamino)phenyl)vinyl]benzoic acid had no detectable water solubility at pH 7.4, and measurements were restricted to (E)-3-[2-(4-(dimethylamino)phenyl)vinyl]benzoic acid (DS3CO) and (E)-2-[2-(4-(dimethylamino)phenyl)vinyl]benzoic acid (DS2CO). While DS3CO and DS2CO were substrates, they also inhibited FMO1 turnover. DS3CO was the more effective inhibitor, and at 2 mM it inhibited FMO1 and microsomal-catalyzed oxidation of methimazole (N-methyl-2-mercaptoimidazole) by 80-90%. Kinetic studies indicated that the aminostilbene carboxylates were noncompetitive with both the xenobiotic substrate, methimazole, and NADPH. However, inhibition constants calculated from double reciprocal plots of velocity vs NADPH were K(i)(comp) 130 and 150 microM for DS3CO and DS2CO, respectively, whereas the uncompetitive Ki's were 10-15 times higher, which suggests that inhibition of NADPH binding may be primarily responsible for inhibition of FMO1 by the aminostilbene carboxylates. This model is also consistent with inhibition of cyclohexanone monooxygenase, a bacterial analog of FMO. DS3CO and DS2CO were again noncompetitive with methimazole but primarily competitive with NADPH. The aminostilbene carboxylates had no detectable effects on activity of pig or rat liver NADPH-cytochrome P450 reductase, which suggests that they are not nonspecific flavoprotein antagonists.
Clement
B
B
Pharmazeutisches Institut, Christian-Albrechts-Universität Kiel, Germany.
Weide
M
M
Ziegler
D M
DM
D000818:Animals; D001565:Benzoates / Q000494:pharmacology*; D002462:Cell Membrane / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects* / Q000378:metabolism; D010101:Oxygen Consumption / Q000187:drug effects; D010105:Oxygenases / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D051381:Rats; D017207:Rats, Sprague-Dawley; D013237:Stereoisomerism; D013267:Stilbenes / Q000138:chemical synthesis* / Q000494:pharmacology* / Q000633:toxicity; D013379:Substrate Specificity
D001565:Benzoates; D004791:Enzyme Inhibitors; D013267:Stilbenes; D010105:Oxygenases; C020066:dimethylaniline monooxygenase (N-oxide forming)