Mixed-function monooxygenase

Hydroxy vitamin D pools ia the blood and is transported on DBF to the kidney, where further hydroxylation takes place at C-1 or C-24 ia response to calcium levels. l-Hydroxylation occurs primarily ia the kidney mitochondria and is cataly2ed by a mixed-function monooxygenase with a specific cytochrome P-450 (52,179,180). 1 a- and 24-Hydroxylation of 25-hydroxycholecalciferol has also been shown to take place ia the placenta of pregnant mammals and ia bone cells, as well as ia the epidermis. Low phosphate levels also stimulate 1,25-dihydtoxycholecalciferol production, which ia turn stimulates intestinal calcium as well as phosphoms absorption. It also mobilizes these minerals from bone and decreases their kidney excretion. Together with PTH, calcitriol also stimulates renal reabsorption of the calcium and phosphoms by the proximal tubules (51,141,181—183). 

Ahokas, J.T. Metabolism of 2,5-diphenyloxazole (PP0) by trout liver microsomal mixed function monooxygenase. Res. Commun. Chem. Pathol. Pharmacol. (1976) 13, 1+39-1+1+7. 

Ahokas, J. T., KSrki, N. T., Oikari, A., and Soivio, A. Mixed-function monooxygenase of fish as an indicator of pollution of aquatic environment by industrial effluent. Bull. Environ. Cont. Toxicol. (1976) 16 270-274. 

Although it is beyond the scope of this chapter to discuss the other oxygen-utilizing enzymes for which there are no crystal structures, it should be noted that mixed-function monooxygenases and oxidases ap-... 

Lambert GH, Kotake AN, Schoeller D. The C02 breath tests as monitors of the cytochrome P450 dependent mixed function monooxygenase system. Prog Clin Biol Res 1983 135 119-145. 

Salicylate is determined in blood serum using immobilized salicylate hydroxylase electrodes (243-245). This enzyme is a mixed function monooxygenase that converts salicylate to catechol in the presence of NAD(P)H and molecular oxygen ... 

Vitamin D-25-hydroxylase of rat liver is a mixed function monooxygenase as demonstrated by the incorporation of 2 into the 25 position of vitamin D The reaction is inhibited by metyrapone, and C0 02 of 9 1. The CO inhibition is released with white light. Vitamin D inhibits aminopyrine demethylase. Thus all evidence currently available suggests this to be a cytochrome P-450 system of the type shown in Fig. 3. Phenobarbital does not induce the vitamin D-25-hydroxylase despite claims to the contrary (T. Madhok and H. F. DeLuca, unpublished re-... 

Cinnamic acid 4-hydroxyla a (EC 1.14.13.11) a mixed function monooxygenase, present in plants, which catalyses an early reaction in Flavonoid (see) biosynthesis, i.e. the insertion of an atom of oxygen into cinnamic acid to form 4-hydroxydnnamic acid (4-coumaric acid) with concomitant oxidation of one molecule of NADPH. The enzyme is a cytochrome P4S0 system associated with the microsom fraction, and is specific for the trans isomer of cinnamic acid. During the hydroxylation, hydrogen at position 4 (experimentally tritium in position 4) is retained, i.e. there is an NIH shift (see). In vitro, a thiol, e.g. 2-mercaptoethanol, is required for activity. (P.R.Rich C.J.Lamb Ear. J.Biochem. 72 (1977) 353-360)... 

Fig. 5. Simplified model of the mixed-function monooxygenase system.

The fact that the same enzyme system can be involved both in lethal synthesis and detoxication can again lead to interactions between different dietary anutrients or drugs. Thus a number of antioxidants (butylated hydroxytoluene, butylated hydroxyanisole, ethoxy-quin ) induce mixed-function monooxygenase, a process which may be considered an adaptive response. As a consequence of this induction, the toxicity of co-administered compounds may be enhanced or reduced, depending on whether the increased activity of the enzymes leads to increased production of toxic or inactive metabolites. In addition, the activity of these pathways is affected by nutritional status, which can further modify the toxic response. The problems of defining a unique no-untoward-effect level are obvious. 

Monooxygenases (Mixed-Function Oxidases, Hydroxylases) Incorporate Only One Atom of Molecular Oxygen Into the Substrate... 

Cytochrome P450 is considered the most versatile biocatalyst known. The actual reaction mechanism is complex and has been briefly described previously (Figure 11-6). It has been shown by the use of that one atom of oxygen enters R—OH and one atom enters water. This dual fate of the oxygen accounts for the former naming of monooxygenases as mixed-function oxidases. The reaction catalyzed by cytochrome P450 can also be represented as follows ... 

In mammals the cytochrome P-U50 mediated monooxygenase or mixed function oxidase system involved in the elimination of lipophilic environmental contaminants and other foreign compounds, has been implicated in the carcinogen activation process. There are several distinct variants of cytochrome P-U50 in mammalian tissues and there may be more than one form of this ubiquitous cytochrome also in fish. The significance of this lies in the fact that different forms of cytochrome P-U50 result in different metabolite patterns, which in turn may reflect on the carcinogenicity or toxicity of compounds being metabolized. 

Phase I renders xenobiotics more polar. Oxidation is the most important process of phase I. It is carried out in the endoplasmic reticulum in many tissues by monooxygenases that contain cytochrome P450 (P450-dependent mixed function oxidases) as electron carrier. These enzymes have evolved in the past billion years in response to plant secondary metabolites. There are a number of P450 gene families. 

For foreign compounds, the majority of oxidation reactions are catalyzed by monooxygenase enzymes, which are part of the mixed function oxidase (MFO) system and are found in the SER (and also known as microsomal enzymes). Other enzymes involved in the oxidation of xenobiotics are found in other organelles such as the mitochondria and the cytosol. Thus, amine oxidases located in the mitochondria, xanthine oxidase, alcohol dehydrogenase in the cytosol, the prostaglandin synthetase system, and various other peroxidases may all be involved in the oxidation of foreign compounds. 

Figure 4.2 The catalytic cycle of the cytochrome(s) P-450 monooxygenase (MFO) system. aFor explanation of step 4, see text. Abbreviation Cyp, cytochromes P-450 MFO, mixed function oxidase. Source From Ref. 1.

Phenylalanine hydroxylase (also called phenylala-nine-4-monooxygenase) is one of a general class of enzymes called mixed-function oxidases (see Box 21-1), all of which catalyze simultaneous hydroxylation of a substrate by an oxygen atom of 02 and reduction of the other oxygen atom to H20. Phenylalanine hydroxylase... 

FIGURE 21-37 Ring closure converts linear squalene to the condensed steroid nucleus. The first step in this sequence is catalyzed by a mixed-function oxidase (a monooxygenase), for which the cosubstrate is NADPH. The product is an epoxide, which in the next step is cyclized to the steroid nucleus. The final product of these reactions in animal cells is cholesterol in other organisms, slightly different sterols are produced, as shown. 

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