Thioredoxin reductase from the malaria mosquito Anopheles gambiae -: Comparisons with the orthologous enzymes of Plasmodium falciparum and the human host
Thioredoxin reductase from the malaria mosquito Anopheles gambiae -: Comparisons with the orthologous enzymes of Plasmodium falciparum and the human host
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DOI：
10.1046/j.1432-1033.2003.03812.x
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发表时间：
2003-11-01
期刊：
影响因子：
--
通讯作者：
Müller, HM
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文献类型：
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作者：
Bauer, H;Gromer, S;Müller, HM
冈比亚按蚊是恶性疟原虫疟疾的重要传播媒介。全基因组分析显示，与果蝇一样，A. gambiae，并且在功能上被硫氧还蛋白系统取代。该系统的关键酶是硫氧还蛋白还原酶-1，一种每亚基55.3 kDa的含FAD的同源二聚体蛋白，其催化NADPH + H+ +硫氧还蛋白二硫化物的反应。NADP+ +硫氧还蛋白二硫醇。助理冈比亚trxr基因作为单拷贝位于X染色体上;它代表编码两种胞质变体和一种线粒体变体的三种剪接变体。优势亚型A. gambiae硫氧还蛋白还原酶-1在大肠杆菌中重组表达，并在功能上与分离的野生型酶进行比较，最终产量为U.ml（-1）包装的昆虫细胞。在氧化还原滴定中，底物A。冈比亚硫氧还蛋白-1（K-m = 8.5 μ M，k（cat）= 15.4 s（-1），pH7.4，25 ℃）不能氧化NADPH还原的A。冈比亚硫氧还蛋白还原酶-1转化为完全氧化状态。这表明，与其它二硫键还原酶相比，A.冈比亚硫氧还蛋白还原酶-1在催化期间在四电子还原态和两电子还原态之间振荡。比较了疟疾系统的硫氧还蛋白还原酶。A.冈比亚硫氧还蛋白还原酶-1与其来自人和恶性疟原虫的直向同源物分别具有52%和45%的序列同一性。这三种酶之间的一个主要区别是C-末端氧化还原中心的结构，反映在催化中间体对自氧化的不同抗性上。该中心的相关序列为A.冈比亚硫氧还蛋白还原酶中的Gly-Cys-硒代半胱氨酸-GlyOH和恶性疟原虫酶中的Cys-X-X-X-X-Cys-GlyOH。这些差异提供了一个有趣的方法来设计物种特异性抑制剂。值得注意的是，A.冈比亚硫氧还蛋白还原酶-1不是硒酶，而是含有高度不寻常的氧化还原活性Cys-Cys序列。
The mosquito, Anopheles gambiae, is an important vector of Plasmodium falciparum malaria. Full genome analysis revealed that, as in Drosophila melanogaster, the enzyme glutathione reductase is absent in A. gambiae and functionally substituted by the thioredoxin system. The key enzyme of this system is thioredoxin reductase-1, a homodimeric FAD-containing protein of 55.3 kDa per subunit, which catalyses the reaction NADPH + H+ + thioredoxin disulfide. NADP+ + thioredoxin dithiol. The A. gambiae trxr gene is located on chromosome X as a single copy; it represents three splice variants coding for two cytosolic and one mitochondrial variant. The predominant isoform, A. gambiae thioredoxin reductase-1, was recombinantly expressed in Escherichia coli and functionally compared with the wild-type enzyme isolated in a final yield of 1.4 U.ml(-1) of packed insect cells. In redox titrations, the substrate A. gambiae thioredoxin-1 (K-m = 8.5 muM, k(cat) = 15.4 s(-1) at pH 7.4 and 25degreesC) was unable to oxidize NADPH-reduced A. gambiae thioredoxin reductase-1 to the fully oxidized state. This indicates that, in contrast to other disulfide reductases, A. gambiae thioredoxin reductase-1 oscillates during catalysis between the four-electron reduced state and a two-electron reduced state. The thioredoxin reductases of the malaria system were compared. A. gambiae thioredoxin reductase-1 shares 52% and 45% sequence identity with its orthologues from humans and P. falciparum, respectively. A major difference among the three enzymes is the structure of the C-terminal redox centre, reflected in the varying resistance of catalytic intermediates to autoxidation. The relevant sequences of this centre are Thr-Cys-Cys-SerOH in A. gambiae thioredoxin reductase, Gly-Cys-selenocysteine-GlyOH in human thioredoxin reductase, and Cys-X-X-X-X-Cys-GlyOH in the P. falciparum enzyme. These differences offer an interesting approach to the design of species-specific inhibitors. Notably, A. gambiae thioredoxin reductase-1 is not a selenoenzyme but instead contains a highly unusual redoxactive Cys-Cys sequence.