Vitamin hydroxylases

Parathyroid hormone, a polypeptide of 83 amino acid residues, mol wt 9500, is produced by the parathyroid glands. Release of PTH is activated by a decrease of blood Ca " to below normal levels. PTH increases blood Ca " concentration by increasing resorption of bone, renal reabsorption of calcium, and absorption of calcium from the intestine. A cAMP mechanism is also involved in the action of PTH. Parathyroid hormone induces formation of 1-hydroxylase in the kidney, requited in formation of the active metabolite of vitamin D (see Vitamins, vitamin d). 

Metabolites of vitamin D, eg, cholecalciferol (CC), are essential in maintaining the appropriate blood level of Ca ". The active metabolite, 1,25-dihydroxycholecalciferol (1,25-DHCC), is synthesized in two steps. In the fiver, CC is hydroxylated to 25-hydroxycholecalciferol (25-HCC) which, in combination with a globulin carrier, is transported to the kidney where it is converted to 1,25-DHCC. This step, which requites 1-hydroxylase formation, induced by PTH, may be the controlling step in regulating Ca " concentration. The sites of action of 1,25-DHCC are the bones and the intestine. Formation of 1,25-DHCC is limited by an inactivation process, ie, conversion of 25-HCC to 24,25-DHCC, catalyzed by 24-hydroxylase. 

Although it is being found that vitamin D metaboUtes play a role ia many different biological functions, metaboHsm primarily occurs to maintain the calcium homeostasis of the body. When calcium semm levels fall below the normal range, 1 a,25-dihydroxy-vitainin is made when calcium levels are at or above this level, 24,25-dihydroxycholecalciferol is made, and 1 a-hydroxylase activity is discontiaued. The calcium homeostasis mechanism iavolves a hypocalcemic stimulus, which iaduces the secretion of parathyroid hormone. This causes phosphate diuresis ia the kidney, which stimulates the 1 a-hydroxylase activity and causes the hydroxylation of 25-hydroxy-vitamin D to 1 a,25-dihydroxycholecalciferol. Parathyroid hormone and 1,25-dihydroxycholecalciferol act at the bone site cooperatively to stimulate calcium mobilization from the bone (see Hormones). Calcium blood levels are also iafluenced by the effects of the metaboUte on intestinal absorption and renal resorption. 

FIGURE 6.17 Hydroxylation of proUne residnes is catalyzed by prolyl hydroxylase. The reaction requires -ketoglntarate and ascorbic acid (vitamin C). 

Scurvy results from a dietary vitamin C deficiency and involves the inability to form collagen fibrils properly. This is the result of reduced activity of prolyl hydroxylase, which is vitamin C-dependent, as previously noted. Scurvy leads to lesions in the skin and blood vessels, and, in its advanced stages, it can lead to grotesque disfiguration and eventual death. Although rare in the modern world, it was a disease well known to sea-faring explorers in earlier times who did not appreciate the importance of fresh fruits and vegetables in the diet. 

CYP24 is a 25-hydroxyvitamin D3 24-hydroxylase that degrades vitamin D metabolites. 

CYP27A1 catalyzes the side chain oxidation (27-hydroxylation) in bile acid biosynthesis. Because bile acid synthesis is the only elimination pathway for cholesterol, mutations in the CYP27A1 gene lead to abnormal deposition of cholesterol and cholestanol in various tissues. This sterol storage disorder is known as cerebrotendinous xanthomatosis. CYP27B1 is the 1-alpha hydroxylase of vitamin D3 that converts it to the active vitamin form. The function of CYP27C1 is not yet known. 

A number of iron-containing, ascorbate-requiring hydroxylases share a common reaction mechanism in which hydroxylation of the substrate is linked to decarboxylation of a-ketoglutarate (Figure 28-11). Many of these enzymes are involved in the modification of precursor proteins. Proline and lysine hydroxylases are required for the postsynthetic modification of procollagen to collagen, and prohne hydroxylase is also required in formation of osteocalcin and the Clq component of complement. Aspartate P-hydroxylase is required for the postsynthetic modification of the precursor of protein C, the vitamin K-dependent protease which hydrolyzes activated factor V in the blood clotting cascade. TrimethyUysine and y-butyrobetaine hydroxylases are required for the synthesis of carnitine. 

Stimulating activation of vitamin D by 1-a-hydroxylase to cal-citriol (1,25-dihydroxyvitmin D3) to promote calcium absorption in the GI tract and increased calcium mobilization from bone... 

Lead has been shown to decrease circulating levels of the active form of vitamin D (1,25-dihydroxy-vitamin D) in children. The conversion of vitamin D to this active hormonal form takes place via hydroxylation to 25-hydroxyvitamin D in the liver, followed by 1-hydroxylation in the mitochondria of the renal tubule by a complex cytochrome P-450 (heme-containing) system (Mahaffey et al. 1982 Rosen and Chesney 1983). Comparisons of the serum 1,25-dihydroxyvitamin D levels in children with blood lead levels of 33 g/dL with those in children with severe renal insufficiency (Rosen et al. 1980) and in children with an inborn error of vitamin D metabolism in which the 1-hydroxylase system or component... 

The final mechanism of action of PTH involves the activation of vitamin D3 through the stimulation of la-hydroxylase in the kidney. In the gastrointestinal tract, vitamin D3 is essential for the absorption of calcium. Enhanced absorption of calcium from dietary sources serves to further increase the concentration of calcium in the blood. Many foods, in particular, dairy products, which are rich in calcium, are fortified with vitamin D. The release of PTH from the parathyroid glands is regulated by plasma calcium levels through negative feedback. A decrease in the level of calcium in the blood stimulates the secretion of PTH and an increase in the calcium level in the blood inhibits it. 

The first step is catalysed by the tetrahydrobiopterin-dependent enzyme tyrosine hydroxylase (tyrosine 3-monooxygenase), which is regulated by end-product feedback is the rate controlling step in this pathway. A second hydroxylation reaction, that of dopamine to noradrenaline (norepinephrine) (dopamine [3 oxygenase) requires ascorbate (vitamin C). The final reaction is the conversion of noradrenaline (norepinephrine) to adrenaline (epinephrine). This is a methylation step catalysed by phenylethanolamine-jV-methyl transferase (PNMT) in which S-adenosylmethionine (SAM) acts as the methyl group donor. Contrast this with catechol-O-methyl transferase (COMT) which takes part in catecholamine degradation (Section 4.6). 

The two hydroxylase enzymes can also utilize the plant-derived steroid, ergocalci-ferol, (vitamin D2) as a substrate. The final product is biologically active and so food manufacturers often fortify their products with ergocalciferol to prevent the occurrence of vitamin D deficiency and consequent rickets in childhood or osteomalacia in adults. 

There seems to be no metabolic control exerted on hepatic 25-hydroxylase and so all of the available cholecalciferol is converted. Hydroxylation in the kidney however is an important control point being regulated by PTH, and indirectly therefore by calcium and phosphate concentrations. Stimulation of la-hydroxylase by PTH is via a cyclic AMP (cAMP) -dependent mechanism and longer-term regulation of the activity of this enzyme is via induction mediated by other hormones such as oestrogens, cortisol and growth hormone. Typically, the plasma concentration of 1,25 dihydroxy vitamin D is in the range 20-60 ng/1, that is approximately 1000-times lower than that of its precursor. 

When 1,25 DHCC provision is adequate or when plasma calcium concentration is above approximately 2.20mmol/l, la hydroxylase activity is suppressed and 25-hydroxy vitamin D3 is converted by 24-hydroxylase into 24,25 dihydroxy vitamin D3 a metabolite whose true role is uncertain but one which seems to have little if any physiological activity. Renal 24-hydroxylase does have a role to play in the deactivation of 1,25 dihydroxy vitamin D the major metabolite of the vitamin being 1,24,25 trihydroxy vitamin D. 

Vitamin D deficiency may also occur through inadequate dietary intake, gut (poor absorption), renal disease (1-hydroxylase deficiency or failure to reclaim calcium from the glomerular filtrate), or liver disease (25-hydroxylase deficiency). The slightly low haemoglobin concentration and pale stained (hypochromic) red cells suggested a coincident mild iron deficiency. 

Danan JL, Delorme AC, Mathieu H. 1982. Presence of 25-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D3 24-hydroxylase in vitamin D target cells of rat yolk sac. J Biol Chem 257 10715-10721. 

Eyles DW, Smith S, Kinobe R, Hewison M, McGrath JJ. 2005. Distribution of the vitamin D receptor and 1 alpha-hydroxylase in human brain. J Chem Neuroanat 29 21-30. 

Matkovits T, Christakos S. 1995. Variable in vivo regulation of rat vitamin D-dependent genes (osteopontin, Ca, Mg-adenosine triphosphatase, and 25-hydroxyvitamin D3 24-hydroxylase) implications for differing mechanisms of regulation and involvement of multiple factors. Endocrinology 136 3971-3982. 

Ren S, Nguyen L, Wu S, Encinas C, Adams JS, et al. 2005. Alternative splicing of vitamin D-24-hydroxylase a novel mechanism for the regulation of extrarenal 1,25-dihydrox-yvitamin D synthesis. J Biol Chem 280 20604-20611. 

Zehnder D, Bland R, Williams MC, McNinch RW, Howieison M. 2001. Extrarenal expression of 25-hydroxy-vitamin d(3)-l alpha-hydroxylase. J Clin Endocrinol Metab... 

L-Asp hydroxylated at the /3-carbon to generate 7a ro-3-hydroxyl-L-aspartic acid has so far only been detected in cinnamycin and the duramycins, lantibiotics produced by actinomycetes. " This modification has also been found in mammalian proteins, such as the vitamin K-dependent protein C, and the epidermal growth factor (EGF)-like domain in human plasma factor IX. Both bovine and human aspartyl-/3-hydroxylases have been purified and characterized and their in vitro hydroxylation activity has been shown using proteins... 

Selected prolines and lysines are hydroxylated by prolyl and lysyl hydroxylases. These enzymes, located in the RER, require ascorbate (vitamin C), deficiency of which produces scurvy. 

Hypocalcemia (below-normal blood calcium) stimulates release of parathyroid hormone (PTH), which in turn binds to receptors on cells of the renal proximal tubules. The receptors are coupled through cAMP to activation of a la-hydroxylase important for the final, rate-hmiting step in the conversion of vitamin D to 1,25-DHCC (dihydroxycholecalciferol or caldtriol). 

Genetic defldency of the la-hydroxylase (vitamin D-resistant rickets)... 

Answer E. The patient has many signs of scurvy from a vitamin C deficiency. The diet, which contains no fruits or vegetables, provides little vitamin C, Prolyl hydroxylase requires vitamin C, and in the absence of hydroxylation, the collagen a-chains do not form stable, mature collagen. The anemia may be due to poor iron absorption in the absence of ascorbate. 

Vitamin C, ascorbic acid, is a coenzyme for the enzyme prolyl hydroxylase. The action of this enzyme is critical for the formation of normal collagen, a key component of structural and connective tissues. 

Ascorbic acid a coenzyme for prolyl hydroxylase the preventive and cure for scurvy also known as vitamin C. 

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