The Arabidopsis plastidial thioredoxins -: New functions and new insights into specificity
The Arabidopsis plastidial thioredoxins -: New functions and new insights into specificity
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DOI：
10.1074/jbc.m302077200
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发表时间：
2003-06-27
影响因子：
4.8
通讯作者：
Miginiac-Maslow, M
中科院分区：
文献类型：
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作者：
Collin, V;Issakidis-Bourguet, E;Miginiac-Maslow, M
拟南芥基因组测序显示，该植物含有大量硫氧还蛋白（Trx）同工型，这是一种参与硫-二硫交换的蛋白质。在序列比较的基础上，鉴定出7个推定的叶绿体Trxs，其中4个属于m型，2个属于f型，1个属于新的x型。在目前的工作中，这些异构体被生产和纯化为重组蛋白，不含其假定的传递肽。用两种已知的叶绿体硫氧还蛋白靶标：nadp -苹果酸脱氢酶和果糖-1,6-二磷酸酶以及叶绿体2-Cys过氧化物还蛋白检测了它们的活性。本研究证实了果糖-二磷酸酶对Trx f的严格特异性，揭示了一些Trx不能激活nadp -苹果酸脱氢酶，并表明新的x型是过氧化氧还蛋白最有效的底物，而对另外两个靶点无活性。这表明这种异构体可能专门参与抗氧化应激。三维模型表明，其中一种m型Trx，即Trx m3，对三种靶标都没有活性，其活性位点周围呈现带负电荷的表面。绿色荧光蛋白方法证实了这些Trxs的质体定位。
The sequencing of the genome of Arabidopsis thaliana revealed that this plant contained numerous isoforms of thioredoxin (Trx), a protein involved in thiol-disulfide exchanges. On the basis of sequence comparison, seven putative chloroplastic Trxs have been identified, four belonging to the m-type, two belonging to the f-type, and one belonging to a new x-type. In the present work, these isoforms were produced and purified as recombinant proteins without their putative transit peptides. Their activities were tested with two known chloroplast thioredoxin targets: NADP-malate dehydrogenase and fructose-1,6-bisphosphatase and also with a chloroplastic 2-Cys peroxiredoxin. The study confirms the strict specificity of fructose-bisphosphatase for Trx f, reveals that some Trxs are unable to activate NADP-malate dehydrogenase, and shows that the new x-type is the most efficient substrate for peroxiredoxin while being inactive toward the two other targets. This suggests that this isoform might be specifically involved in resistance against oxidative stress. Three-dimensional modeling shows that one of the m-type Trxs, Trx m3, which has no activity with any of the three targets, exhibits a negatively charged surface surrounding the active site. A green fluorescent protein approach confirms the plastidial localization of these Trxs.