Effect of mutations at Lys250, Arg251, and Lys253 of cytochrome P450 1A2 on the catalytic activities and the bindings of bifunctional axial ligands.
Effect of mutations at Lys250, Arg251, and Lys253 of cytochrome P450 1A2 on the catalytic activities and the bindings of bifunctional axial ligands.
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DOI：
10.1016/0003-9861(92)90113-b
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发表时间：
1992-10
影响因子：
3.9
通讯作者：
A. Krainev;T. Shimizu;K. Hiroya;M. Hatano
中科院分区：
文献类型：
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作者：
A. Krainev;T. Shimizu;K. Hiroya;M. Hatano
一些真核细胞色素P450(P450)含有一系列离子氨基酸，对应于P450 1A2序列中的Lys250、Arg251和Lys253残基。为了了解这些离子氨基酸在P450催化功能中的作用，从酵母表达系统中获得了P450 1A2的3个单一突变体Lys250Leu、Arg251Leu和Lys253Leu。与野生型相比，Arg251Leu突变体在P450重组体系催化的甲氧基和乙氧基间苯二酚脱烷基化反应中的周转次数显著增加了6倍。Lys250Leu和Lys253Leu突变体的营业额也比野生型高三到四倍。这些催化活性被吡啶衍生物、含氮的轴向配体与P450血红素竞争抑制。根据这些发现，结合其他光谱数据，推测Lys250、Arg251和Lys253的离子位置可能位于该酶的底物识别位置和/或轴向配体通道附近。
Some eukaryotic cytochromes P450 (P450s) have a series of ionic amino acids, corresponding to Lys250, Arg251, and Lys253 residues in the P450 1A2 sequence. To understand the roles of those ionic amino acids in the catalytic function of P450, three single mutants, Lys250Leu, Arg251Leu, and Lys253Leu of P450 1A2 were obtained from yeast (Saccharomyces cerevisiae) expression system. Turnover numbers of the Arg251Leu mutant in dealkylation reactions of methoxy- and ethoxyresorufin catalyzed by the P450 reconstituted system were remarkably increased by sixfold compared to those of the wild type. The Lys250Leu and Lys253Leu mutants also showed turnover numbers higher than those of the wild type by three- to fourfold. Those catalytic activities were inhibited competitively by pyridine derivatives, nitrogenous axial ligands to the P450 heme. From those findings together with other spectral data, it was suggested that the ionic site of Lys250, Arg251, and Lys253 may be somehow located near the substrate recognition site and/or near the axial-ligand access channel of this enzyme.