The Escherichia coli heat shock protease HtrA participates in defense against oxidative stress
The Escherichia coli heat shock protease HtrA participates in defense against oxidative stress
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DOI：
10.1007/s004380051092
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发表时间：
1999-09
期刊：
影响因子：
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通讯作者：
J. Skórko-Glonek;D. Zurawa;E. Kuczwara;M. Wozniak;Z. Wypych;B. Lipińska
中科院分区：
文献类型：
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作者：
J. Skórko-Glonek;D. Zurawa;E. Kuczwara;M. Wozniak;Z. Wypych;B. Lipińska
丝氨酸蛋白酶 HtrA (DegP) 是细胞在高温下生存所必需的，是一种外周膜蛋白，位于大肠杆菌内膜的周质侧，生化和遗传学证据表明，HtrA 的生理作用是降解热休克期间细胞包膜中形成的变性蛋白质。本研究的目的是查明 HtrA 蛋白酶是否有助于保护细胞免受氧化应激。我们比较了各种氧化剂对 htrAmutant 细胞的影响和对野生型细菌的影响，发现 htrAmutation 并没有增加对过氧化氢或百草枯的敏感性，但使细胞对亚铁 [Fe(II)] 离子极其敏感，而亚铁离子会增强蛋白质的氧化。用亚铁离子处理导致细胞膜部分中的蛋白质羰基水平比周质和细胞质中的蛋白质羰基水平增加更大。相对于野生型细胞，htrA突变体中铁诱导的膜蛋白氧化增强。与主要作用于细胞可溶部分的衍生物（4OH-TEMPO）相比，位于细胞膜中的硝基氧抗氧化剂（A-TEMPO）可以更有效地减轻铁对 htrAmutant 生长的抑制。用氢过氧化异丙苯处理后，对htrAmutant生长的抑制更为明显，氢过氧化异丙苯可分配到膜中，而用氢过氧化叔丁基处理则更明显，氢过氧化叔丁基主要在细胞溶胶中形成自由基。亚铁离子和氢过氧化枯烯均诱导 HtrA 的合成，但过氧化氢、百草枯或叔丁基氢过氧化物不诱导 HtrA 的合成。我们的结果表明，HtrA 在防御氧化休克中发挥作用，并支持 HtrA 参与细胞膜中氧化损伤蛋白质（尤其是与细胞膜相关的蛋白质）降解的假设。
The serine protease HtrA (DegP), which is indispensable for cell survival at elevated temperatures, is a peripheral membrane protein, localized on the periplasmic side of the inner membrane inEscherichia coli, and the biochemical and genetic evidence indicates that the physiological role of HtrA is to degrade denatured proteins formed in the cellular envelope during heat shock. The aim of this study was to find out if the HtrA protease contributes to protection of the cell against oxidative stress. We compared the influence of various oxidizing agents onhtrAmutant cells with their effects on wild-type bacteria, and found that thehtrAmutation did not increase sensitivity to hydrogen peroxide or paraquat but made the cell extremely sensitive to ferrous [Fe(II)] ions, which are known to enhance oxidation of proteins. Treatment with ferrous ions caused a larger increase in the level of protein carbonyl groups in the membrane fraction of the cell than in the periplasm and cytoplasm. Iron-induced oxidation of membrane proteins was enhanced in thehtrAmutant relative to wild-type cells. Inhibition of the growth of thehtrAmutant by iron could be alleviated more efficiently by a nitroxide antioxidant that localizes in the membranes (A-TEMPO) than by a derivative (4OH-TEMPO) that acts mainly in the soluble fraction of the cell. Inhibition of the growth of thehtrAmutant was more pronounced following treatment with cumene hydroperoxide, which partitions into membranes, than witht-butyl hydroperoxide, which forms radical mainly in the cytosol. Both ferrous ions and cumene hydroperoxide, but not hydrogen peroxide, paraquat ort-butyl hydroperoxide, induced synthesis of HtrA. Our results show that HtrA plays a role in defense against oxidative shock and support the hypothesis that HtrA participates in the degradation of oxidatively damaged proteins localized in the cell envelope, especially those associated with the membranes.