Reconstitution of liver monooxygenase system in solution from cytochrome P-450 and NADPH-specific flavoprotein monomers.
Microsomal monooxygenase system was reconstituted in the presence of non-ionic detergent Emulgen 913 from cytochrome P-450 and NADPH-specific flavoprotein isolated from phenobarbital-induced rabbit liver microsomes. At Emulgen 913 concentration of 0.05 g/l mixed complex between flavoprotein and cytochrome was formed with 5: 5 protein molar ratio and molecular weight of 700 kD. The 2-hour incubation of the enzymes with 0.25 g/l Emulgen 913 at 4 degrees C was accompanied by dissociation of protein oligomers to monomers. The reconstituted systems containing flavoprotein and cytochrome as mixed complexes or monomers were able to catalyze NADPH-dependent cytochrome P-450 reduction and benzphetamine N-demethylation. Taking into consideration the effective concentrations of the enzymes the apparent second order rate constants of these reactions with monomers were 100 times those with complexes.
D000818:Animals; D003577:Cytochrome P-450 Enzyme System; D005420:Flavoproteins / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D008297:Male; D008433:Mathematics; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D009249:NADP / Q000378:metabolism*; D017137:Nonoxynol; D010089:Oxidoreductases, N-Demethylating / Q000378:metabolism; D010105:Oxygenases / Q000378:metabolism*; D010634:Phenobarbital / Q000494:pharmacology; D011092:Polyethylene Glycols / Q000494:pharmacology; D011817:Rabbits
D005420:Flavoproteins; D007527:Isoenzymes; D017137:Nonoxynol; D011092:Polyethylene Glycols; D009249:NADP; D003577:Cytochrome P-450 Enzyme System; D010105:Oxygenases; C019781:benzphetamine N-demethylase; D010089:Oxidoreductases, N-Demethylating; D010634:Phenobarbital