Tomato cytochrome P450CYP734A7 functions in brassinosteroid catabolism
Tomato cytochrome P450CYP734A7 functions in brassinosteroid catabolism
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DOI：
10.1016/j.phytochem.2006.05.042
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发表时间：
2006-09-01
期刊：
影响因子：
3.8
通讯作者：
Mizutani, Masaharu
中科院分区：
文献类型：
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作者：
Ohnishi, Toshiyuki;Nomura, Takahito;Mizutani, Masaharu
一些细胞色素P450单加氧酶(P450)在油菜素类固醇(BR)的生物合成和BR分解代谢中催化必要的氧化反应；然而，关于参与BR分解代谢途径的P450的信息有限。在这里，我们报道了番茄中两个P450mRNAs的特性，分别是CYP734A7和CYP734A8。这些P450与拟南芥CYP734A1/BAS1(以前的CYP72B1)有很高的同源性，后者通过C-26羟基化来失活BRS。结构性过表达CYP734A7的转基因烟草表现出与BR缺乏症相似的极端矮化表型。气相色谱-质谱法对转基因植株内源BRs的定量分析表明，与野生型植株相比，转基因植株中的蓖麻酮和6-脱氧卡斯特酮的含量显著降低。用重组细胞色素P734A7测定底物结合光谱，测得蓖麻酮、油菜素内酯和6-脱氧卡斯特酮的解离常数分别为6.7、12和12微米。在体外实验中，细胞色素P734A7被证实能将蓖麻酮代谢成26-羟基蓖麻甾酮。此外，还将28-去甲孕酮和油菜素内酯转化为羟基化产物。外源生物活性BRS对番茄幼苗细胞色素P734A7和细胞色素P734A8基因的表达有上调作用。这些结果表明，CYP734A7是BRS的C-26羟基酶，可能参与番茄BR的分解代谢。细胞色素P734A亚家族在多种植物中的存在表明，这些蛋白对BRs的氧化失活是植物中普遍存在的现象。(C)2006爱思唯尔有限公司。保留所有权利。
Several cytochrome P450 monooxygenases (P450s) catalyze essential oxidative reactions in brassinosteroid (BR) biosynthesis as well as in BR catabolism; however, only limited information exists on the P450s involved in the BR catabolic pathway. Here, we report the characterization of two P450 mRNAs, CYP734A7 and CYP734A8, from Lycopersicon esculentum. These P450s show high homology with Arabidopsis CYP734A1/BAS1 (formerly CYP72B1), which inactivates BRs via C-26 hydroxylation. Transgenic tobacco plants that constitutively overexpressed CYP734A7 showed an extreme dwarf phenotype similar to BR deficiency. Quantitative gas chromatography-mass spectrometry analysis of endogenous BRs in the transgenic plants showed that the levels of castasterone and 6-deoxocastasterone significantly decreased in comparison with those in wild-type plants. By measuring the Type I substrate-binding spectra using recombinant CYP734A7, the dissociation constants for castasterone, brassinolide, and 6-deoxocastasterone were determined to be 6.7, 12, and 12 mu M, respectively. In an in vitro assay, CYP734A7 was confirmed to metabolize castasterone to 26-hydroxycastasterone. In addition, 28-norcastasterone and brassinolide were converted to the hydroxylated products. The expression of CYP734A7 and CYP734A8 genes in tomato seedlings was upregulated by exogenous application of bioactive BRs. These results indicated that CYP734A7 is a C-26 hydroxylase of BRs and is likely involved in BR catabolism in tomato. The presence of the CYP734A subfamily in various plant species suggests that oxidative inactivation of BRs by these proteins is a widespread phenomenon in plants. (c) 2006 Elsevier Ltd. All rights reserved.