Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1993 Sep 15;90(18):8668–8672. doi: 10.1073/pnas.90.18.8668

Transfected human liver cytochrome P-450 hydroxylates vitamin D analogs at different side-chain positions.

Y D Guo 1, S Strugnell 1, D W Back 1, G Jones 1
PMCID: PMC47419  PMID: 7690968

Abstract

A full-length cDNA for the human liver mitochondrial cytochrome P-450 CYP27 was cloned from a human hepatoma HepG2 cDNA library and then subcloned into the mammalian expression vector pSG5. When CYP27 cDNA was transfected into COS-1 transformed monkey kidney cells along with adrenodoxin cDNA, transfected cells revealed a 10- to 20-fold higher vitamin D3-25-hydroxylase activity than nontransfected cells. Transfected cells were capable of 25-hydroxylation of vitamin D3, 1 alpha-hydroxyvitamin D3 and 1 alpha-hydroxydihydrotachysterol3. In each case they also showed the ability to 26(27)-hydroxylate the cholesterol-like (D3) side chain. The relative rates of 25- and 26(27)-hydroxylation of 1 alpha-hydroxyvitamin D3 approximately mimicked the ratio of products observed in HepG2 cells. Vitamin D2 and 1 alpha-hydroxyvitamin D2, both with the ergosterol-like side chain, were 24- and 26(27)-hydroxylated by CYP27. The rate of side-chain 24-, 25-, or 26(27)-hydroxylation was greater for 1 alpha-hydroxylated vitamin D analogs than for their nonhydroxylated counterparts. We conclude that CYP27 is capable of 24-, 25-, and 26(27)-hydroxylation of vitamin D analogs and that the nature of products is partially dictated by the side chain of the substrate. This work has revealed that the cytochrome P-450 CYP27 may be important in the metabolism of vitamin D analogs used as drugs.

Full text

PDF
8668

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Andersson S., Holmberg I., Wikvall K. 25-hydroxylation of C27-steroids and vitamin D3 by a constitutive cytochrome P-450 from rat liver microsomes. J Biol Chem. 1983 Jun 10;258(11):6777–6781. [PubMed] [Google Scholar]
  2. Bhattacharyya M. H., DeLuca H. F. The regulation of rat liver calciferol-25-hydroxylase. J Biol Chem. 1973 May 10;248(9):2969–2973. [PubMed] [Google Scholar]
  3. Björkhem I., Holmberg I., Oftebro H., Pedersen J. I. Properties of a reconstituted vitamin D3 25-hydroxylase from rat liver mitochondria. J Biol Chem. 1980 Jun 10;255(11):5244–5249. [PubMed] [Google Scholar]
  4. Cali J. J., Hsieh C. L., Francke U., Russell D. W. Mutations in the bile acid biosynthetic enzyme sterol 27-hydroxylase underlie cerebrotendinous xanthomatosis. J Biol Chem. 1991 Apr 25;266(12):7779–7783. [PMC free article] [PubMed] [Google Scholar]
  5. Cali J. J., Russell D. W. Characterization of human sterol 27-hydroxylase. A mitochondrial cytochrome P-450 that catalyzes multiple oxidation reaction in bile acid biosynthesis. J Biol Chem. 1991 Apr 25;266(12):7774–7778. [PubMed] [Google Scholar]
  6. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  7. Fukushima M., Suzuki Y., Tohira Y., Nishii Y., Suzuki M. 25-Hydroxylation of 1alpha-hydroxyvitamin D3 in vivo and in perfused rat liver. FEBS Lett. 1976 Jun 1;65(2):211–214. doi: 10.1016/0014-5793(76)80482-6. [DOI] [PubMed] [Google Scholar]
  8. Holmberg I., Berlin T., Ewerth S., Björkhem I. 25-Hydroxylase activity in subcellular fractions from human liver. Evidence for different rates of mitochondrial hydroxylation of vitamin D2 and D3. Scand J Clin Lab Invest. 1986 Dec;46(8):785–790. doi: 10.3109/00365518609084051. [DOI] [PubMed] [Google Scholar]
  9. Horst R. L., Koszewski N. J., Reinhardt T. A. 1 alpha-hydroxylation of 24-hydroxyvitamin D2 represents a minor physiological pathway for the activation of vitamin D2 in mammals. Biochemistry. 1990 Jan 16;29(2):578–582. doi: 10.1021/bi00454a035. [DOI] [PubMed] [Google Scholar]
  10. Jones G. A new pathway of 25-hydroxyvitamin D3 metabolism. Methods Enzymol. 1986;123:141–154. doi: 10.1016/s0076-6879(86)23017-7. [DOI] [PubMed] [Google Scholar]
  11. Jones G., Schnoes H. K., Levan L., Deluca H. F. Isolation and identification of 24-hydroxyvitamin D2 and 24,25-dihydroxyvitamin D2. Arch Biochem Biophys. 1980 Jul;202(2):450–457. doi: 10.1016/0003-9861(80)90449-x. [DOI] [PubMed] [Google Scholar]
  12. Nebert D. W., Nelson D. R., Coon M. J., Estabrook R. W., Feyereisen R., Fujii-Kuriyama Y., Gonzalez F. J., Guengerich F. P., Gunsalus I. C., Johnson E. F. The P450 superfamily: update on new sequences, gene mapping, and recommended nomenclature. DNA Cell Biol. 1991 Jan-Feb;10(1):1–14. doi: 10.1089/dna.1991.10.1. [DOI] [PubMed] [Google Scholar]
  13. Qaw F., Calverley M. J., Schroeder N. J., Trafford D. J., Makin H. L., Jones G. In vivo metabolism of the vitamin D analog, dihydrotachysterol. Evidence for formation of 1 alpha,25- and 1 beta,25-dihydroxy-dihydrotachysterol metabolites and studies of their biological activity. J Biol Chem. 1993 Jan 5;268(1):282–292. [PubMed] [Google Scholar]
  14. Roise D., Schatz G. Mitochondrial presequences. J Biol Chem. 1988 Apr 5;263(10):4509–4511. [PubMed] [Google Scholar]
  15. Saarem K., Bergseth S., Oftebro H., Pedersen J. I. Subcellular localization of vitamin D3 25-hydroxylase in human liver. J Biol Chem. 1984 Sep 10;259(17):10936–10940. [PubMed] [Google Scholar]
  16. Su P., Rennert H., Shayiq R. M., Yamamoto R., Zheng Y. M., Addya S., Strauss J. F., 3rd, Avadhani N. G. A cDNA encoding a rat mitochondrial cytochrome P450 catalyzing both the 26-hydroxylation of cholesterol and 25-hydroxylation of vitamin D3: gonadotropic regulation of the cognate mRNA in ovaries. DNA Cell Biol. 1990 Nov;9(9):657–667. doi: 10.1089/dna.1990.9.657. [DOI] [PubMed] [Google Scholar]
  17. Tam S. P., Strugnell S., Deeley R. G., Jones G. 25-Hydroxylation of vitamin D3 in the human hepatoma cell lines Hep G2 and Hep 3B. J Lipid Res. 1988 Dec;29(12):1637–1642. [PubMed] [Google Scholar]
  18. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Usui E., Noshiro M., Okuda K. Molecular cloning of cDNA for vitamin D3 25-hydroxylase from rat liver mitochondria. FEBS Lett. 1990 Mar 12;262(1):135–138. doi: 10.1016/0014-5793(90)80172-f. [DOI] [PubMed] [Google Scholar]
  20. Wichmann J., Schnoes H. K., DeLuca H. F. Isolation of identification of 24(R)-hydroxyvitamin D3 from chicks give large doses of vitamin D3. Biochemistry. 1981 Apr 14;20(8):2350–2353. doi: 10.1021/bi00511a043. [DOI] [PubMed] [Google Scholar]
  21. Yoon P. S., Deluca H. F. Resolution and reconstitution of soluble components of rat liver microsomal vitamin D3-25-hydroxylase. Arch Biochem Biophys. 1980 Sep;203(2):529–541. doi: 10.1016/0003-9861(80)90210-6. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES