Quantifying changes in the thiol redox proteome upon oxidative stress in vivo
Quantifying changes in the thiol redox proteome upon oxidative stress in vivo
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DOI：
10.1073/pnas.0707723105
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发表时间：
2008-06-17
影响因子：
11.1
通讯作者：
Jakob, Ursula
中科院分区：
文献类型：
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作者：
Leichert, Lars I.;Gehrke, Florian;Jakob, Ursula
抗微生物水平的活性氧（ROS）是由哺乳动物宿主防御杀死入侵细菌和限制细菌定植产生的。体内活性氧的一个主要靶点是蛋白质的巯基。我们开发了一种称为OxICAT的定量硫醇捕获技术，用于鉴定体内过氧化氢（H2O2）和次氯酸盐（NaOCI）应激的生理重要靶蛋白。OxICAT允许在单个实验中对数百种不同蛋白质的氧化硫醇修饰进行精确定量。它还可以识别受影响的蛋白质并确定其氧化还原敏感的半胱氨酸。利用这种技术，我们鉴定了一组具有显著（30-90%）氧化修饰硫醇基团的大肠杆菌蛋白，它们似乎对H2O2或NaOCI胁迫特别敏感。这些结果表明，单个氧化剂在体内针对不同的蛋白质。条件必需的大肠杆菌基因编码三分之一的氧化还原敏感蛋白，这一发现可能解释氧化应激治疗的抑菌效果。我们确定了一组氧化还原调节蛋白，保护大肠杆菌免受氧化应激条件。这些实验表明，OxICAT可用于各种不同的细胞类型和生物体，是鉴定、量化和监测体内氧化硫醇修饰的有力工具。
Antimicrobial levels of reactive oxygen species (ROS) are produced by the mammalian host defense to kill invading bacteria and limit bacterial colonization. One main in vivo target of ROS is the thiol group of proteins. We have developed a quantitative thiol trapping technique termed OxICAT to identify physiologically important target proteins of hydrogen peroxide (H2O2) and hypochlorite (NaOCI) stress in vivo. OxICAT allows the precise quantification of oxidative thiol modifications in hundreds of different proteins in a single experiment. It also identifies the affected proteins and defines their redox-sensitive cysteine(s). Using this technique, we identified a group of Escherichia coli proteins with significantly (30-90%) oxidatively modified thiol groups, which appear to be specifically sensitive to either H2O2 or NaOCI stress. These results indicate that individual oxidants target distinct proteins in vivo. Conditionally essential E. coli genes encode one-third of redox-sensitive proteins, a finding that might explain the bacteriostatic effect of oxidative stress treatment. We identified a select group of redox-regulated proteins, which protect E. coli against oxidative stress conditions. These experiments illustrate that OxICAT, which can be used in a variety of different cell types and organisms, is a powerful tool to identify, quantify, and monitor oxidative thiol modifications in vivo.