Reconstitution of liver monooxygenase system in solution from cytochrome P-450 and NADPH-specific flavoprotein...

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.