Functional analysis of Arabidopsis thaliana isoforms of the Mg-chelatase CHLI subunit
Functional analysis of Arabidopsis thaliana isoforms of the Mg-chelatase CHLI subunit
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DOI：
10.1039/b802604c
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发表时间：
2008-10-01
影响因子：
3.1
通讯作者：
Masuda, Tatsuru
中科院分区：
文献类型：
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作者：
Kobayashi, Koichi;Mochizuki, Nobuyoshi;Masuda, Tatsuru
叶绿素生物合成的第一步是由一个由CHLI、CHILD和CHLH组成的mg螯合酶催化。拟南芥有两个CHLI亚型CHLI1和CHLI2，它们具有相似的表达谱，但有研究表明CHLI2在mg -螯合酶复合体中的功能有限。最近，我们发现拟南芥CHLI I是一个atp酶和叶绿体硫氧还蛋白的靶标。在这里，我们证明CHLI2也具有ATP酶活性，但比CHLI1具有更低的V-max和更高的K-m （ATP）。我们证实了CHLI2中硫氧还蛋白依赖的二硫键还原和硫醇对其atp酶活性的调节。然后，我们使用CHLI2 T-DNA敲除系检查了CHLI2的生理贡献。虽然纯合chli2突变体的可见表型与野生型几乎相当，但突变体的叶绿素积累明显减少。此外,cs / cs;Chli2 / Chli2双突变体几乎是白化的。从cs/cs分离出来的后代有三种表型；， CHLI2/ CHLI2亲本：cs状淡绿色。黄色和几乎白化的比例约为1:2:7 .7。PCR分析证实，在纯合子背景下，chli2突变为半显性。这些结果表明，虽然CHLI2在叶绿素生物合成中发挥有限的作用，但该亚基肯定有助于mg螯合酶复合物的组装。
The first step of chlorophyll biosynthesis is catalyzed by a Mg-chelatase composed of the subunits CHLI, CHILD and CHLH. Mg-chelatase requires ATP hydrolysis that can be attributed to CHILL Arabidopsis has two CHLI isoforms, CHLI1 and CHLI2, that have similar expression profiles, but it has been suggested that CHLI2 has limited function in the Mg-chelatase complex. Recently, we showed that Arabidopsis CHLI I is an ATPase and a target of chloroplast thioredoxin. Here, we demonstrate that CHLI2 also has ATPase activity but with a lower V-max and higher K-m (ATP) than CHLI1. We confirmed the thioredoxin-dependent reduction of a disulfide bond in CHLI2 and thiol-modulation of its ATPase activity. We then examined the physiological contribution of CHLI2 using a chli2 T-DNA knockout line. Although visible phenotype of homozygous chli2 mutants was almost comparable to wild type, the mutant accumulated significantly less chlorophyll. Furthermore, cs/cs; chli2/chli2 double mutants were almost albino. There were three phenotypes among progenies segregated from the cs/cs;, CHLI2/chli2 parent: cs-like pale green. yellow, and almost albino were obtained in the approximate ratio of 1:2:0.7. PCR analysis confirmed that the chli2 mutation is semidominant on a homozygous cs background. These results reveal that although CHLI2 plays a limited role in chlorophyll biosynthesis, this subunit certainly contributes to the assembly of the Mg-chelatase complex.