Adrenal cortex, iron

In mitochondria (Fig. lb), the electron acceptor protein is also a flavoprotein termed NADPH-adrenodoxin reductase (MW 50 kDa) because it was discovered in the adrenal cortex and because it donates its electrons not directly to the P450 but to the smaller redox protein adrenodoxin (MW 12.5 kDa). The two iron-sulphur clusters of this protein serve as electron shuttle between the flavoprotein and the mitochondrial P450. 

Figure 11-6. Cytochrome P450 hydroxylase cycle in microsomes. The system shown is typical of steroid hydroxylases of the adrenal cortex. Liver microsomal cytochrome P450 hydroxylase does not require the iron-sulfur protein FejSj. Carbon monoxide (CO) inhibits the indicated step.

Fritz, J., Anderson, R., Fee, J., Palmer, G, Sands, R.H., Tsibris, J.C.M., Gunsalus, I.C., Orme-Johnson, W.H., and Beinert, H. 1971. The iron electron-nuclear double resonance (ENDOR) of two-iron ferredoxins from spinach, parsley, pig adrenal cortex and Pseudomonas putida. Biochimica et Biophysica Acta 253 110-133. 

Adrenodoxin. Adrenodoxin is the only iron-sulfur protein which has been isolated from mammals. This protein from mitochondria of bovine adrenal cortex was purified almost simultaneously by Kimura and Suzuki (32) and Omura et al. (33). It has a molecular weight of 12,638 (34) and the oxidized form of the protein shows maximal absorbances at 415 and 453 nm. Adrenodoxin acts as an electron carrier protein in the enzyme system required for steroid hydroxylation in adrenal mitochondria. In this system, electron transfer is involved with three proteins cytochrome P. gQ, adrenodoxin and a flavoprotein. Reduced NADP gives an electron to Tne flavoprotein which passes the electron to adrenodoxin. Finally, reduced adrenodoxin transfers the electron to cytochrome Pas shown in Fig. 3. The mechanism of cytochrome P cq interaction with steroid, oxygen and adrenodoxin in mixed-function oxidase of adrenal cortex mitochondria has been reviewed by Estabrook et al. (35). 

A partial list of physiological functions til at have been determined to be affected by vitamin C deficiencies includes (1) absorption of iron (2) cold tolerance, maintenance of adrenal cortex (3) antioxidant (4) metabolism of tryptophan, phenylalanine, and tyrosine (5) body growth (6) wound healing (7) synthesis of polysaccharides and collagen (8) formation of cartilage, dentine, bone, and teeth and (9) maintenance of capillaries. 

Omura, T., E. Sanders, D. Y. Cooper, O. Rosenthal, and R. W. Estrabrook Isolation of a non-heme iron protein of adrenal cortex functional as a TPNH-flavoprotein-cytochrome P 50 reductase for hydroxylation reactions. In Non-Heme Iron Proteins Role in Energy Conversion, A. San Pietro, ed., Antioch Press, Yellow Springs, Ohio, pp. 401—412 (1965). 

Figure 17.8 Typical hydroxylation reaction involving cytochrome P450. Ad, adrenodoxin. The substrate is a steroid hormone biosynthesis intermediate. The location of the reaction is in the mitochondria of adrenal cortex. (Reproduced by permission from Bezkorovainy A. Biochemistry of Nonheme Iron. New York Plenum Press, 1980, p. 380.)...

The purification procedure for the pig testis non-heme iron protein has been published elsewhere (28, 43). The procedure is similar to Procedure B . The isolation of testis mitochondrial fraction, Procedure A , is not recommended due to the low yield of the mitochondria and loss of the non-heme iron protein. The yield of the non-heme iron protein from testis was very much lower than the yield of adrenodoxin from adrenal cortex. This may indicate that testis hydroxylation is less active than adrenal hydroxylation. 

Cytochromes are used in the conversion of cholesterol to the steroid hormones. Iliese hormones include aldosterone, cortisol, and the sex hormones. Synthesis of aldosterone, for example, occurs in the mitochondria of the adrenal cortex. One of the steps in aidostero ne synthesis is a hydroxylation that is cataly .ed by a complex of adrenodoxin reductase (FAD-containing protein), adrenodoxin (nonheme iron protein), and cytochrome P450 (heme protein). 

Omura, T., E. Sanders, R.W. Estabrook, D.Y. Cooper, and O. Rosenthal (1966). Isolation from adrenal cortex of a nonheme iron protein and a flavoprotein functional as a reduced triphosphopy-ridine nucleotide-cytochrome P-450 reductase. Arch. Biochem. Biophys. 117, 660-673. 

Hypophysectomy, which produces an atrophy of the adrenal cortex, has been reported to inhibit [122, 298] and to enhance [425] the egg-white reaction or to have no effect on the egg-white [401] and dextran [396] reactions. More recently, Gabbiani and Bocskor [199] reported that hypophysectomy reduces the sensitivity of rats to iron-dextran and that normal sensitivity is restored by the transfusion of plasma from an animal with an intact pituitary. This suggests that a plasma substance is produced in the pituitary gland which has a permissive action on the anaphylactoid reaction. This report has yet to be confirmed. 

W.R. Dunham, et al., 2-Iron ferredoxins in spinach, parsley, pig adrenal cortex, Azotobacter vinelandii, and Clostridium pasteur-ianum—studies by magnetic field Mossbauer spectroscopy. 2, Biochim. Biophys. Acta 1971, 253(1), 134. 

The existence of mitochondrial cytochrome P450 in adrenal cortex was reported originally by Harding et al (1964) and confirmed by subsequent studies. Adrenal cortical mitochondria catalyze a number of NADPH- and molecular-oxygen-dependent hydroxylation reactions that contribute to the biosynthesis of corticosteroids. The enzyme system for 11 jS-hydroxyla-tion has been isolated, and a successful reconstitution of its activity has been achieved by the interaction of three proteins, namely an NADPH-dependent flavoprotein (adrenodoxin reductase), an iron-sulfur protein (adrenodoxin), and the heme protein (cytochrome P450) that serves as the terminal oxidase for the electron transport system from NADPH to oxygen (Wang and... 

Why is vitamin C used so extensively as an electron donor There are two main reasons. First, vitamin C is very soluble in water, so it can be concentrated in confined spaces surrounded by membranes (which are made of lipids impermeable to vitamin C). The synthesis of noradrenaline from dopamine, for example, takes place in small membrane-bounded spaces, or vesicles, within cells of the cortex of the adrenal glands. The vitamin C concentration inside these vesicles reaches about 100 times that of blood plasma. As vitamin C is consumed by the enzyme dopamine mono-oxygenase, electrons are passed across the vesicle membrane (via an iron-containing protein, cytochrome b65l), to regenerate vitamin C within the vesicles. Thus, for periods of days or weeks, the intracellular vitamin C needed for physiological tasks can be insulated from changes in plasma levels caused by variations in diet, and maintained at the ideal levels for a particular reaction. 

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