| 虎杖查尔酮合酶PcCHS1基因的克隆与功能分析 |
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| 作者姓名： | 李星 王红 |
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| 作者单位： | 中国科学院研究生院, 北京 100049 |
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| 基金项目： | 国家自然科学基金(61173098)和中国科学院"十二五"基础前沿专项(KSCX2-EW-J-29)资助 |
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| 摘 要： | 从中药虎杖中通过RACE等方法克隆到一个查尔酮合酶基因的全长cDNA,命名为PcCHS1.该cDNA全长1182 bp,编码一个含393个氨基酸的蛋白质.体外酶促活性研究表明,重组PcCHS1在pH 7～8时催化形成查尔酮为其单一产物,在pH 9时除催化形成查尔酮外,还产生一定量的苯亚甲基丙酮.对PcCHS1第216位和第333位氨基酸进行了定点突变研究,结果表明这些位点对PcCHS1的体外酶促活性影响较大.
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| 关 键 词： | 虎杖 查尔酮合酶 定点突变 酶活性 |
| 收稿时间： | 2012-03-29 |
| 修稿时间： | 2012-04-13 |
Cloning and characterization of PcCHS1 from Polygonum cuspidatum |
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| Authors: | LI Xing WANG Hong |
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| Institution: | College of life science, Graduate University, Chinese Academy of Sciences, Beijing 100049, China |
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| Abstract: | A full length cDNA encoding a type Ⅲ PKS (PcCHS1) was isolated from Polygonum cuspidatum Sieb. et Zucc. Functional and enzymatic assays biochemically confirmed that PcCHS1 was a chalcone synthase. When incubated with p-coumaroyl-CoA and malonyl-CoA at pH 7-8, PcCHS1 catalyzed the formation of chalcone as the major product. However, at pH 9 both p-hydroxybenzalacetone and chalcone were detected. Site-directed mutagenesis of Phe216 into Leu caused reduction of enzyme activity. Another mutation N333K performed activity of releasing some common derailment products, CTAL and BNY, of many CHSs in vitro. |
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| Keywords: | Polygonum cuspidatum" target="_blank">Polygonum cuspidatum')">Polygonum cuspidatum chalcone synthase site-directed mutagenesis enzyme activity |
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