title: Peroxidase activity of liver microsomal vitamin D 25-hydroxylase and cytochrome P450 1A2 catalyzes 25-hydroxylation of vitamin D3 and oxidation of dopamine to aminochrome.
abstract: Purified liver microsomal vitamin D 25-hydroxylase, a cytochrome P450, catalyzes 25-hydroxylation of vitamin D3 in the absence of NADPH and NADPH-cytochrome P450 reductase by using t-butyl hydroperoxide as electron donors. The rate of 25-hydroxylation was approximately the same when NADPH/ NADPH-cytochrome P450 reductase or t-butyl hydroperoxide was used as electron donor. The Km value for vitamin D3 in the presence of t-butyl hydroperoxide was found to be 30 microM. The rates of 25-hydroxylation of 1 alpha-hydroxyvitamin D3 and 5 beta-cholestane-3 alpha, 7 alpha-diol catalyzed by 25-hydroxylase were significantly higher when the reaction proceeded in the presence of NADPH/NADPH-cytochrome P450 reductase than in the presence of t-butyl hydroperoxide. Other liver microsomal cytochrome P450 forms such as taurochenodeoxycholic acid 6 alpha-hydroxylase and cytochromes P450 1A2 and 2B4 did not catalyze 25-hydroxylation of vitamin D3 in the presence of NADPH/NADPH-cytochrome P450 reductase or t-butyl hydroperoxide. The peroxidase activity of the 25-hydroxylase also catalyzed oxidation of dopamine to aminochrome. A linear correlation between increase in aminochrome formation and increase in the amount of 25-hydroxylase was observed in the oxidation of dopamine. The Km values for dopamine and t-butyl hydroperoxide were 8.6 microM and 1 mM when 25-hydroxylase catalyzes the formation of aminochrome in the presence of t-butyl hydroperoxide. Oxidation of dopamine to aminochrome catalyzed by the peroxidase activity of cytochrome P-450 1A2 was observed in the presence of t-butyl hydroperoxide. A linear correlation between formation of aminochrome and the amount of cytochrome P450 1A2 was found.
authors.affiliation: Department of Pharmaceutical Biosciences, University of Uppsala, Sweden.
authors.forename: J
authors.initials: J
authors.lastname: Segura-Aguilar
mesh_terms: D000818:Animals; D002762:Cholecalciferol / Q000378:metabolism*; D053493:Cholestanetriol 26-Monooxygenase; D019388:Cytochrome P-450 CYP1A2 / Q000378:metabolism*; D004298:Dopamine / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism; D045563:Indolequinones; D007211:Indoles / Q000378:metabolism*; D008862:Microsomes, Liver / Q000201:enzymology*; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D011817:Rabbits; D013250:Steroid Hydroxylases; D013552:Swine
chemical_list: D045563:Indolequinones; D007211:Indoles; D002762:Cholecalciferol; C091584:aminochrome 1; D009249:NADP; D006861:Hydrogen Peroxide; D013250:Steroid Hydroxylases; D019388:Cytochrome P-450 CYP1A2; D053493:Cholestanetriol 26-Monooxygenase; D004298:Dopamine