灰色链霉菌细胞色素P450SU-1将维生素D3转化为1α,25-二羟基维生素D3 。
Conversion of vitamin D3 to 1alpha,25-dihydroxyvitamin D3 by Streptomyces griseolus cytochrome P450SU-1.
作者信息
Sawada Natsumi, Sakaki Toshiyuki, Yoneda Sachiyo, Kusudo Tatsuya, Shinkyo Raku, Ohta Miho, Inouye Kuniyo
机构信息
Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
出版信息
Biochem Biophys Res Commun. 2004 Jul 16;320(1):156-64. doi: 10.1016/j.bbrc.2004.05.140.
Streptomyces griseolus cytochrome P450SU-1 (CYP105A1) was expressed in Escherichia coli at a level of 1.0 micromol/L culture and purified with a specific content of 18.0 nmol/mg protein. Enzymatic studies revealed that CYP105A1 had 25-hydroxylation activity towards vitamin D2 and vitamin D3. Surprisingly, CYP105A1 also showed 1alpha-hydroxylation activity towards 25(OH)D3. As mammalian mitochondrial CYP27A1 catalyzes a similar two-step hydroxylation towards vitamin D3, the enzymatic properties of CYP105A1 were compared with those of human CYP27A1. The major metabolite of vitamin D2 by CYP105A1 was 25(OH)D2, while the major metabolites by CYP27A1 were both 24(OH)D2 and 27(OH)D2. These results suggest that CYP105A1 recognizes both vitamin D2 and vitamin D3 in a similar manner, while CYP27A1 does not. The Km values of CYP105A1 for vitamin D2 25-hydroxylation, vitamin D3 25-hydroxylation, and 25-hydroxyvitamin D3 1alpha-hydroxylation were 0.59, 0.54, and 0.91 microM, respectively, suggesting a high affinity of CYP105A1 for these substrates.
灰色链霉菌细胞色素P450SU-1（CYP105A1）在大肠杆菌中表达，表达水平为1.0微摩尔/升培养物，并以18.0纳摩尔/毫克蛋白质的特定含量进行纯化。酶学研究表明，CYP105A1对维生素D2和维生素D3具有25-羟化活性。令人惊讶的是，CYP105A1对25(OH)D3也表现出1α-羟化活性。由于哺乳动物线粒体CYP27A1对维生素D3催化类似的两步羟化反应，因此将CYP105A1的酶学性质与人类CYP27A1的进行了比较。CYP105A1催化维生素D2的主要代谢产物是25(OH)D2，而CYP27A1催化的主要代谢产物是24(OH)D2和27(OH)D2。这些结果表明，CYP105A1以相似的方式识别维生素D2和维生素D3，而CYP27A1则不然。CYP105A1对维生素D2 25-羟化、维生素D3 25-羟化和25-羟基维生素D3 1α-羟化的Km值分别为0.59、0.54和0.91微摩尔，表明CYP105A1对这些底物具有高亲和力。
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