کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1991664 | 1541024 | 2012 | 9 صفحه PDF | دانلود رایگان |
![عکس صفحه اول مقاله: Metabolism of cholesterol, vitamin D3 and 20-hydroxyvitamin D3 incorporated into phospholipid vesicles by human CYP27A1 Metabolism of cholesterol, vitamin D3 and 20-hydroxyvitamin D3 incorporated into phospholipid vesicles by human CYP27A1](/preview/png/1991664.png)
CYP27A1 is a mitochondrial cytochrome P450 which can hydroxylate vitamin D3 and cholesterol at carbons 25 and 26, respectively. The product of vitamin D3 metabolism, 25-hydroxyvitamin D3, is the precursor to the biologically active hormone, 1α,25-dihydroxyvitamin D3. CYP27A1 is attached to the inner mitochondrial membrane and substrates appear to reach the active site through the membrane phase. We have therefore examined the ability of bacterially expressed and purified CYP27A1 to metabolize substrates incorporated into phospholipid vesicles which resemble the inner mitochondrial membrane. We also examined the ability of CYP27A1 to metabolize 20-hydroxyvitamin D3 (20(OH)D3), a novel non-calcemic form of vitamin D derived from CYP11A1 action on vitamin D3 which has anti-proliferative activity on keratinocytes, leukemic and myeloid cells. CYP27A1 displayed high catalytic activity towards cholesterol with a turnover number (kcat) of 9.8 min−1 and Km of 0.49 mol/mol phospholipid (510 μM phospholipid). The Km value of vitamin D3 was similar for that of cholesterol, but the kcat was 4.5-fold lower. 20(OH)D3 was metabolized by CYP27A1 to two major products with a kcat/Km that was 2.5-fold higher than that for vitamin D3, suggesting that 20(OH)D3 could effectively compete with vitamin D3 for catalysis. NMR and mass spectrometric analyses revealed that the two major products were 20,25-dihydroxyvitamin D3 and 20,26-dihydroxyvitamin D3, in almost equal proportions. Thus, the presence of the 20-hydroxyl group on the vitamin D3 side chain enables it to be metabolized more efficiently than vitamin D3, with carbon 26 in addition to carbon 25 becoming a major site of hydroxylation. Our study reports the highest kcat for the 25-hydroxylation of vitamin D3 by any human cytochrome P450 suggesting that CYP27A1 might be an important contributor to the synthesis of 25-hydroxyvitamin D3, particularly in tissues where it is highly expressed.
► CYP27A1 metabolizes cholesterol and vitamin D3 in phospholipid vesicles.
► Comparable Km values observed for the metabolism of vitamin D3 and cholesterol.
► CYP27A1 metabolizes CYP11A1-derived 20-hydroxyvitamin D3 to two major products.
► Products identified as 20,25-dihydroxyvitamin D3 and 20,26-dihydroxyvitamin D3.
► 20-Hydroxyvitamin D3 is metabolized more efficiently than vitamin D3.
Journal: The Journal of Steroid Biochemistry and Molecular Biology - Volume 129, Issues 3–5, April 2012, Pages 163–171