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The role of chlorophyll a oxygenase in the regulation of chlorophyll a/chlorophyll b ratio

叶绿素a加氧酶在叶绿素a/叶绿素b比例调节中的作用

基本信息

批准号：
10440239
负责人：
TANAKA Ayumi
金额：
$ 7.04万
依托单位：
HOKKAIDO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

项目摘要

Chlorophyll b is an ubiquitous accessory pigment in land plants, green algae and prochlorophytes. Its biosynthesis plays a key role in the adaptation to various light environments. We isolated 6 chlorophyll b-less mutants by insertional mutagenesis and isolated a gene responsible for chlorophyll b synthesis whose sequence is similar to those of methyl monooxygenases, and contains putative binding domains for a [2Fe-2S] Rieske center and for a mononuclear iron. These results demonstrate that chlorophyll a oxygenase (CAO) is involved in the chlorophyll b formation. In order to know the reaction mechanisms, we expressed the CAO in E.coli and carried out the enzymatic studies. The recombinant enzyme produced in E.coli catalyses an unusual two-step oxygenase reaction and converted chlorophyllide a to chlorophyllide b.The light-harvesting efficiency of a photosystem is thought to be largely dependent on its photosynthetic antenna size. It has been suggested that antenna size is controlled by the biosynthesis of chlorophyll b. To verify this hypothesis, we overexpressed the enzyme for chlorophyll b biosynthesis (CAO) in Arabidopsis thaliana by transforming the plant with cDNA for CAO under the control of the 35S cauliflower mosaic virus promoter. In the early greening phase, when the intrinsic CAO expression is very low, the chlorophyll a-to-b ratio was drastically decreased from 28 to 7.3, indicating that enhancement of chlorophyll b biosynthesis had been successfully achieved. We also found with full-green rosette leaves of transgenic plants that the chlorophyll a-to-b ratio was reduced from 2.85 to 2.65 and that the ratio of the peripheral light-harvesting complexes to the core antenna complex increased by 20. These results indicated that the antenna size increased in transgenic plants, suggesting that CAO regulates antenna size by controlling chlorophyll b synthesis.

叶绿素B是陆生植物、绿色藻类和原绿藻中普遍存在的辅助色素。它的生物合成在适应各种光环境中起着关键作用。我们分离出6个叶绿素b-少突变体的插入诱变和分离的基因负责叶绿素B的合成，其序列是类似的甲基单加氧酶，并含有推定的结合域的[2Fe-2S] Rieske中心和单核铁。这些结果表明，叶绿素a加氧酶（CAO）参与叶绿素B的形成。为了了解反应机理，我们在大肠杆菌中表达了CAO，并进行了酶促研究。在大肠杆菌中产生的重组酶催化不寻常的两步加氧酶反应，并将叶绿素a转化为叶绿素B。光系统的光捕获效率被认为在很大程度上取决于其光合天线的大小。有人认为，触角的大小受叶绿素B的生物合成控制。为了验证这一假设，我们在拟南芥中过表达的叶绿素B生物合成（CAO）的酶转化植物的cDNA为CAO的35 S花椰菜花叶病毒启动子的控制下。在早期绿化阶段，当内在CAO表达非常低时，叶绿素a与B的比率从28急剧降低到7.3，表明叶绿素B生物合成的增强已经成功实现。我们还发现，转基因植物的全绿色莲座叶的叶绿素a到B的比例从2.85降低到2.65，并且外围捕光复合体与核心天线复合体的比例增加了20。这些结果表明，在转基因植株中，触角的大小增加，表明CAO通过控制叶绿素B的合成来调节触角的大小。