Sequential hydroxylation of vitamin D2 by a genetically engineered CYP105A1.
Sequential hydroxylation of vitamin D2 by a genetically engineered CYP105A1.
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基因工程 CYP105A1 对维生素 D2 进行连续羟基化。
DOI：
10.1016/j.bbrc.2016.03.139
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发表时间：
2016
期刊：
影响因子：
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通讯作者：
T. Sakaki
中科院分区：
文献类型：
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作者：
K. Hayashi;K. Yasuda;Y. Yogo;T. Takita;K. Yasukawa;M. Ohta;M. Kamakura;S. Ikushiro;T. Sakaki
我们之前的研究表明，CYP105A1 - R73A/R84A和R73V/R84A的双变异体在将维生素D3转化为其生物活性形式1α，25-二羟基维生素D3（1α,25(OH)2D3）方面表现出高水平的活性。在本研究中，我们发现这两种双变异体也能够通过25(OH)D2将维生素D2转化为活性形式，即1α，25-二羟基维生素D2(1α,25(OH)2D2)，而其对25(OH)D2的1α-羟基化活性远低于25(OH)D3。野生型和双变异体的比较表明，氨基酸取代显著增强了对维生素D2的25-和26-羟基化活性。在维生素D2的25-羟基化后，C26的进一步羟基化可能经常发生，而不会从底物结合袋中释放25(OH)D2。因此，CYP105A1的双变异体对产生25,26(OH) 2d2非常有用，25,26(OH) 2d2是人血清中检测到的维生素d2的代谢物之一。
Our previous studies revealed that the double variants of CYP105A1— R73A/R84A and R73V/R84A—show high levels of activity with respect to conversion of vitamin D3to its biologically active form, 1α,25-dihydroxyvitamin D3(1α,25(OH)2D3). In this study, we found that both the double variants were also capable of converting vitamin D2to its active form, that is, 1α,25-dihydroxyvitamin D2(1α,25(OH)2D2), via 25(OH)D2,whereas its 1α-hydroxylation activity toward 25(OH)D2was much lower than that toward 25(OH)D3. Comparison of the wild type and the double variants revealed that the amino acid substitutions remarkably enhanced both 25- and 26-hydroxylation activity toward vitamin D2. After 25-hydroxylation of vitamin D2, further hydroxylation at C26 may occur frequently without the release of 25(OH)D2from the substrate-binding pocket. Thus, the double variants of CYP105A1 are quite useful to produce 25,26(OH)2D2that is one of the metabolites of vitamin D2detected in human serum.