Microsomal hydroxylation system

Lu, A. Y. H. and Levin, W. The resolution and reconstitution of the liver microsomal hydroxylation system. Biochim. 

It should be emphasized here that the electron transfer system consisting of adrenodoxin differs from that of the microsomal hydroxylating system. 

The reaction pathway is given in Fig. 33, and the scheme is comparable to the microsomal hydroxylating system. The reducing equivalents are given by NADH and the putidaredoxin. Putidaredoxin is an intermediate compound between the flavoprotein reduced by NADH and cytochrome P-450. 

Jaffe H, Fujii K, Sengupta M, Guerin H, Epstein SS. In vivo inhibition of mouse liver microsomal hydroxylating systems by methylenedioxyphenyl insecticidal synergists and related compounds. Life Sci 1968 7 1051-62. 

The aromatic hydroxylases or mixed-function oxidases are no exception to the above generalization. For maximum activity they require a transition metal ion and an electron donor such as one of the pyridine or flavin nucleotide coenzymes further, they probably utilize molecular oxygen as the source of the hydroxylic oxygen an example is the liver microsomal hydroxylating system (27). As yet there is no comprehensive explanation to cover the mode of action of these enzymes, for on the one hand there are the specific hydroxylases which catalyze such conversions as L-phenylalanine to L-tyrosine (26) or tryptophan to 5-hydroxytrypto-... 

Ginter, E., Kosinova, A., Hudecova, A., and Mlynarcikova, U., 1984, Parabolic response of hepatic microsomal hydroxylating system and lipids to graded doses of ascorbic acid in guinea pigs on low and high a-tocopherol intake, J. Nutr. 114 485-492. 

A similar series of conversions occurs after the administration of the related amino acid, a-methyl-me/a-tyrosine. leading in this case to the production of L-metaraminol. In some species a microsomal hydroxylating system can also catalyse the introduction of a phenolic hydroxyl group into L-metaraminol or one of its precursors, so that the final product stored in the adrenergic nerves may be a mixture of metaraminol and oc-methylNA (Fig. 10). Again the amount of... 

Phosphatidylcholine was demonstrated to play an essential role in the biological activity of the microsomal hydroxylation system (Lu et al., 1969), functioning in some poorly understood manner in electron transfer from NADPH to cytochrome P-450. Reconstitution studies of the microsomal hydroxylation system from its isolated components have demonstrated the ability of a number of nonionic detergents in low concentration to substitute for phospholipid in the process of benzphetamine N-demethylation (Lu et al., 1974). While restoration of catalytic activity was not complete, it is likely that the natural lipid and the detergents act in a similar manner, perhaps enhancing the interaction of the two proteins of this system. 

In vivo intermediate in mammals by the detection of two of its derivatives,, the glutathione (GSH) conjugate and its further metabolites formed by an initial carbamylation reaction (W) (see below) and 2-chloroacrolein detected in the microsome-NADPH system and derived from the rearrangement-elimination reaction sequence discussed above (6). Sulfallate also yields 2-chloroacrolein in the microsome-NADPH system, presumably by -CH2 hydroxylation (22) on analogy with the metabolism of EPTC shown previously. 

West. S.B. and Lu, A.Y. (1972) Reconstituted liver microsomal enzyme system that hydroxylates drugs, other foreign compounds and endogenous substrates. V. Competition between cytochromes P-450 and P-448 for reductase in 3,4-benzpyrene hydro-xylation. Archives of Biochemistry and Biophysics, 153, 298-303. 

The corresponding reactions of the methyl groups at C-4 on the A ring16" 68 73 are depicted on the left side of Fig. 22-8. Tire 4a methyl group is first hydroxylated by a microsomal (ER) system similar to cytochrome P450 but able to accept electrons from NADH and cytochrome b5 rather than NADPH.173 The two-step oxidation of the resulting alcohol... 

Antipyrine is oxidized through biotransformation, independently of perfusion, predominantly in the micro-somes, and is excreted after hydroxylation and conjugation. After oral administration (15 or 18 mg/kg BW, respectively), the metabolic clearance ability of the liver (metabolic capacity of the microsomal monooxygenase system) can be assessed by computation of the concentration curve and the plasma half-life (after 3 and 24 hours). The serum half-life and plasma clearance are significantly enhanced/decreased, depending on the reduction in liver function. There is a close correlation with the galactose elimination capacity as well as with Quick s value. (58-60, 74, 88)... 

Microsomal hydroxylation at allylic carbon atoms is commonly observed in drug metabolism. An illustrative example of allylic oxidation is given by the psychoactive component of marijuana. J -tetrahydnx annabinol J -THC. This molecule contains three allylic carbon centers (C-7. C-6. and C-3). Allylic hydroxylation occurs cxten.sively at C-7 to yield 7-hydroxy- j -THC as the major pla.sma metabolite in humans. " Pharmacological studies show that this 7-hydroxy metabolite is as active as, or even more active than. J -THC per se and may contribute significantly to the overall central nervous system (CNS) p.sychotomimctic effects of the parent compound. Hydroxylation also occurs to a minor extent at the allylic C-6 position to give both the epimeric ba- and 6/3-hydroxy metabolites. " Metaboli.sm does not occur at C-3, presumably becau.se of steric hindrance. 

In 1957, we reported on the use of the detergent deoxycholate for the solubilization of aminoazo dye N-demethylase, and we also reported an inhibitory effect of CO on aminoazo dye N-demethylase activity (49). In a key study in 1968, Lu Coon described the deoxycholate-dependent solubilization and resolution of a liver microsomal fatty acid >-hydroxylation system into three components by column chromatography, and they were able to reconstitute catalytic activity by combining the three fractions (50). The three fractions were identified as cytochrome P450,... 

Lu AYH, Junk KW, Coon MJ. 1969. Resolution of the cytochome P-450 containing co—hydroxylation system of liver microsomes into three components. J. Biol. Chem. 244 3714-21... 

In this model system, as contrasted with the simple ferric ion reductase activity of the flavoprotein (388), the metal is not the ultimate electron acceptor but presumably serves the dual role of oxygen activation and electron carrier. The reaction may involve superoxide anion since it is inhibited by superoxide dismutase (erthrocuprein) (394) Xanthine plus xanthine oxidase can also serve as electron donor, and this latter model system is also inhibited by superoxide dismutase (395). Superoxide dismutase also inhibits the menadione-mediated NADPH oxidase activity of NADPH-cytochrome P-450 (396) as well as the reconstituted benz-phetamine hydroxylation system (397). The involvement of NADPH-cytochrome P-450 reductase in microsomal lipid peroxidation has been confirmed by the demonstration that the reaction in microsomes is totally inhibited by antibody to the purified reductase (374)- It has been suggested that lipid peroxidation by microsomes requires another component, in addition to the reductase, which takes the place of the ferric ion chelate in the model system (374) ... 

Oxidations via microsomal P450 system Aliphatic hydroxylation (pentobarbital)... 

The nuclear changes involved in the conversion of cholesterol to cholic acid (reduction of the 5,6 double bond, epimerization of the hydroxyl group at carbon-3, and insertion of hydroxyl groups at positions 7 and 12) appear to be carried out by microsomal enzyme systems. Side-chain oxidation is effected by mitochondrial enzymes. 

The co-hydroxylation of fatty acids, which involves the addition of a hydroxyl group at or near the co-terminal carbon, was first shown to be catalyzed by the liver microsomal enzyme system in the 1960s [38], In particular, the CO pigment of CYP was recognized as a constituent of the microsomal mixed function oxidase system that contributes to the co-hydroxylation of steroids [39], Substrates that are susceptible to co-hydroxylation include laurate and AA [38], Early reports showed that CYP enzymes catalyze the CO (C-20) and co-1 (C-19) hydroxy lation of AA [40], In 1990, Faick et al. demonstrated that CYP enzymes also hydroxy late the C-16 (co-4), C-17 (co-3), and C-18 (co-2) carbons of A A [41], Thus, in the presence of nicotinamide adenine dinucleotide phosphate (NADPH) and molecular oxygen, CYPs mediate the hydroxylation of AA to generate a variety of co-terminal HETEs including 16-, 17-, 18-, 19-, and 20-HETR (Fig. 13.2). 

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