Transfected human liver cytochrome P-450 hydroxylates vitamin D analogs at different side-chain positions.
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.
Guo
Y D
YD
Department of Biochemistry, Queen's University, Kingston, ON, Canada.
Strugnell
S
S
Back
D W
DW
Jones
G
G
D000818:Animals; D001483:Base Sequence; D006528:Carcinoma, Hepatocellular; D002460:Cell Line; D002522:Chlorocebus aethiops; D053493:Cholestanetriol 26-Monooxygenase; D002851:Chromatography, High Pressure Liquid; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000378:metabolism*; D006801:Humans; D006900:Hydroxylation; D007668:Kidney; D008113:Liver Neoplasms; D013058:Mass Spectrometry; D008930:Mitochondria, Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D011061:Poly A / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods; D012313:RNA / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D013250:Steroid Hydroxylases / Q000096:biosynthesis / Q000378:metabolism*; D013379:Substrate Specificity; D014162:Transfection; D014407:Tumor Cells, Cultured; D014807:Vitamin D / Q000302:isolation & purification / Q000378:metabolism*
D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014807:Vitamin D; D011061:Poly A; D012313:RNA; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; C506831:CYP27A1 protein, human; D053493:Cholestanetriol 26-Monooxygenase