Regulation of Translation by the Redox State of Elongation Factor G in the Cyanobacterium Synechocystis sp PCC 6803
Regulation of Translation by the Redox State of Elongation Factor G in the Cyanobacterium Synechocystis sp PCC 6803
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DOI：
10.1074/jbc.m109.015131
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发表时间：
2009-07-10
影响因子：
4.8
通讯作者：
Nishiyama, Yoshitaka
中科院分区：
文献类型：
--
作者：
Kojima, Kouji;Motohashi, Ken;Nishiyama, Yoshitaka
延伸因子G（EF-G）是翻译延伸中的关键蛋白质，被鉴定为蓝细菌集胞藻属PCC 6803的翻译机制内活性氧类失活的主要靶标（Kojima，K.，Oshita，M.，Nanjo，Y.，加塞，K.，Tozawa，Y.，林，H.，和Nishiyama，Y.（2007）Mol. Microbiol. 65，936-947）。在本研究中，我们发现，EF-G（Slr 1463）的失活H2 O2是由于两个特定的半胱氨酸残基的氧化和二硫键的形成。这些半胱氨酸残基的丝氨酸残基的取代保护EF-G从失活H2 O2，并允许EF-G介导的翻译系统中的体外翻译，已准备从集胞藻。氧化后的EF-G中的二硫键被硫氧还蛋白还原，还原后的EF-G在体外恢复了翻译活性。蛋白质印迹分析表明，EF-G的氧化形式的水平增加，在强光下的突变体，缺乏NADPH-硫氧还蛋白还原酶，表明EF-G是由硫氧还蛋白在体内减少。这些观察结果表明，翻译机制受到EF-G氧化还原状态的调节，EF-G被活性氧物质氧化，并被硫氧还蛋白还原，硫氧还蛋白是一种由光合电子运输产生的还原信号的递质。
Elongation factor G (EF-G), a key protein in translational elongation, was identified as a primary target of inactivation by reactive oxygen species within the translational machinery of the cyanobacterium Synechocystis sp. PCC 6803 (Kojima, K., Oshita, M., Nanjo, Y., Kasai, K., Tozawa, Y., Hayashi, H., and Nishiyama, Y. (2007) Mol. Microbiol. 65, 936-947). In the present study, we found that inactivation of EF-G (Slr1463) by H2O2 was attributable to the oxidation of two specific cysteine residues and formation of a disulfide bond. Substitution of these cysteine residues by serine residues protected EF-G from inactivation by H2O2 and allowed the EF-G to mediate translation in a translation system in vitro that had been prepared from Synechocystis. The disulfide bond in oxidized EF-G was reduced by thioredoxin, and the resultant reduced form of EF-G regained the activity to mediate translation in vitro. Western blotting analysis showed that levels of the oxidized form of EF-G increased under strong light in a mutant that lacked NADPH-thioredoxin reductase, indicating that EF-G is reduced by thioredoxin in vivo. These observations suggest that the translational machinery is regulated by the redox state of EF-G, which is oxidized by reactive oxygen species and reduced by thioredoxin, a transmitter of reducing signals generated by the photosynthetic transport of electrons.