1. The reconstitution of microsomal redox chains. A comparitive analysis of the effectiveness of membrane self-assembly and template binding of electron carriers.
- Author
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Archakov AI, Bachmanova GI, Devichensky YM, Karuzina II, Zherebkova NS, Alimov GA, Kuznetsova GP, and Karyakin AV
- Subjects
- Animals, Cytochrome P-450 Enzyme System metabolism, Cytochromes metabolism, Electron Transport, Flavoproteins, In Vitro Techniques, Kinetics, Liposomes, Male, Membranes metabolism, Microscopy, Electron, Microsomes ultrastructure, NAD, NADH, NADPH Oxidoreductases metabolism, NADP, Oxidation-Reduction, Protein Binding, Rats, Microsomes metabolism
- Abstract
A comparative analysis was made of the effectiveness of three methods for the reconstitution of microsomal electron-transfer chains, namely, self-assembly, incorporation of electron carriers into liposomes (non-specific template) and incorporation into ;ghosts' of microsomal vesicles (specific template). It was shown that when the ;ghosts' of the microsomal vesicles were used as a specific template extra cytochrome b(5) and NADH-specific flavoprotein were incorporated into them, but cytochrome P-450 and NADPH-specific flavoprotein were not incorporated into the membrane. As a result of the self-assembly and incorporation into liposomes all the electron carriers were present in the reconstituted membrane. Cytochrome P-450 reactivation took place and the inactive form, cytochrome P-420, was converted into the active form, cytochrome P-450. Of the four enzyme hydroxylation systems studied, i.e. NADPH- and NADH-dependent p-hydroxylation of aniline, and NADPH- and NADH-dependent N-demethylation of dimethylaniline, only the NADH-dependent demethylation of dimethylaniline (60% of the initial value) and NADH-dependent p-hydroxylation of aniline (30% of the initial value) were reconstituted by self-assembly. NADPH oxidase and NADH oxidase activities were only properly reconstituted by self-assembly and incorporation into liposomes. In contrast, the NADPH-specific system of peroxidation of unsaturated fatty acids was reconstituted by specific template-binding.
- Published
- 1974
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