Bligh H F, Wolf C R, Smith G, Beggs J D
Institute of Cell and Molecular Biology, Edinburgh University, U.K.
Gene. 1992 Jan 2;110(1):33-9. doi: 10.1016/0378-1119(92)90441-q.
We present a novel strategy for increasing the level of functional mammalian cytochrome P450 (Cyt.P450) and NADPH:cytochrome P450 reductase enzymes produced in yeast. A cDNA encoding the rat P450 reductase was modified by the addition of a sequence coding for the N-terminal region of P450 reductase from Saccharomyces cerevisiae. The addition of this hydrophobic tail greatly increased the apparent stability of the reductase protein produced in S. cerevisiae, as compared to the unmodified rat P450 reductase. When the rat hybrid reductase was produced simultaneously with one of two mammalian Cyt.P450s, the rat CYP2B1 or the human CYP2A6, there was a significant increase in the specific activity of each of the Cyt.P450s. The optimization of this approach and its extrapolation to other organisms should lead to a marked improvement in our ability to study and exploit the P450 system.
我们提出了一种新策略，用于提高酵母中产生的功能性哺乳动物细胞色素P450（Cyt.P450）和NADPH：细胞色素P450还原酶的水平。通过添加编码酿酒酵母P450还原酶N端区域的序列，对编码大鼠P450还原酶的cDNA进行了修饰。与未修饰的大鼠P450还原酶相比，添加这条疏水尾巴极大地提高了酿酒酵母中产生的还原酶蛋白的表观稳定性。当大鼠杂交还原酶与两种哺乳动物Cyt.P450之一（大鼠CYP2B1或人CYP2A6）同时产生时，每种Cyt.P450的比活性都有显著提高。对该方法的优化及其向其他生物体的推广应用，应能显著提高我们研究和利用P450系统的能力。
