Polyphyllins are the major steroidal saponin components of Paris polyphylla, the main source plant of the common Chinese herbal medicine Paridis Rhizoma with strong pharmacological activity and extremely high economic value and great market prospects. However, the production of polyphyllins in plants is limited, and their biosynthesis pathway has not been reported. The downstream hydroxylation step was particularly unclear. To clarify the enzymes and intermediates involved in the downstream steps of polyphyllin biosynthesis, we performed a comparative transcriptome analysis and discovered a cytochrome P450 gene that encodes a protein with monooxygenase activity. Heterologous expression in Saccharomyces cerevisiae demonstrated that it encodes an enzyme that catalyzes the formation of 22(R)-hydroxycholesterol from cholesterol. The relative gene expression measured by RT-PCR and polyphyllin contents measured by HPLC in P. polyphylla roots at different ages confirmed that this gene is involved in polyphyllin biosynthesis. To our best knowledge, this is the first report on the cloning of a CYP450 enzyme gene from the steroidal saponin pathway of higher plants.

中文翻译：

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负责巴黎多叶甾体皂苷生物合成途径中 C-22 羟基化步骤的细胞色素 P450 单加氧酶

多叶素是七叶树的主要甾体皂苷成分，是常见中草药香根草的主要来源植物，药理活性强，经济价值极高，市场前景广阔。然而，植物中多叶素的产量有限，其生物合成途径尚未见报道。下游羟基化步骤特别不清楚。为了阐明参与多叶素生物合成下游步骤的酶和中间体，我们进行了比较转录组分析并发现了一种细胞色素 P450 基因，该基因编码具有单加氧酶活性的蛋白质。Saccharomyces cerevisiae 中的异源表达表明它编码一种酶，该酶催化胆固醇形成 22(R)-羟基胆固醇。RT-PCR测定的相对基因表达和HPLC测定的多叶松根在不同龄期的多叶素含量证实该基因参与多叶素的生物合成。据我们所知，这是从高等植物甾体皂苷途径中克隆 CYP450 酶基因的第一份报告。