【摘要】 异黄酮主要分布于豆科植物中，具有丰富的保健功效。异黄酮合成酶(IFS)是一种膜结合细胞色素P450酶(P450),催化独特的芳环迁移和羟基化。到目前为止，研究得到的植物P450酶晶体结构数量有限。通过培养硒代蛋氨酸取代晶体，利用MAD方法测定了蒺藜苜蓿IFS分辨率为1.9?的晶体结构，并进行了分子对接和突变研究。结合咪唑的IFS结构具有Iα螺旋-环-Iβ螺旋基序，与其他P450酶的螺旋I相对应。与常见的P450结构相比，IFS/咪唑复合物结构包含1个额外的结构域，即γ-结构域。IFS/咪唑复合物结构是一种同源二聚体，其中一个分子的γ-结构域与另一个分子的β-结构域发生作用。血红素基团平面与Iα螺旋-环-Iβ螺旋基序形成约40°的夹角。分子对接和突变研究表明，Trp-128和Asp-300可能在底物结合和识别中发挥重要作用。来自Iα螺旋-环-Iβ螺旋基序的Phe-301、Ser-303和Gly-305可能在芳环迁移中发挥重要作用。这些新颖的结构特征揭示了IFS独特的反应机制，为在豆科作物中进行IFS育种工程奠定了基础。更多还原

【Abstract】 Isoflavones which mainly distributed in leguminous plants have plenty of health benefits.Isoflavone synthase(IFS) is a membrane-associated cytochrome P450 enzyme(CYP450) which carries out the unique aryl-ring migration and hydroxylation. So far, few crystal structures of plant P450s have been obtained. We determined the crystal structure of IFS fromMedicago truncatulaat 1. 9 ? by MAD method using a selenomethionine substituted crystal and conducted molecular docking and mutagenesis study. The structure of IFS complexed with imidazole exhibits the helix Iα-loop-helix Iβ motif which corresponds to helix I of other P450s. Compared with structures of common P450s, IFS/imidazole structure contains an extra domain, i. e., the γ-domain. The structure reveals a homodimer in which the γ-domain of one molecule interacts with the β-domain of another. The plane of heme group makes an angle of approximately 40° with the helix Iα-loop-helix Iβ motif. Molecular docking combined with mutagenesis study suggested that Trp-128 and Asp-300 might play important roles in substrate binding and recognition. Phe-301, Ser-303 and Gly-305 from the helix Iα-loop-helix Iβ motif may play important roles in the aryl-ring migration. These novel structural features reveal insights into the unique reaction mechanism of IFS and provide a basis for engineering IFS in leguminous crops for health purpose.更多还原