Peroxiredoxin 6 as an Anti-oxidant Enzyme

过氧化还原蛋白 6 作为抗氧化酶

基本信息

批准号：
7796689
负责人：
Aron B. FISHER
金额：
$ 35.67万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-05-01 至 2010-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7796689
关键词：
Active Sites Affect Alcohols Alveolar Animals Antioxidants Back Binding Binding Proteins Binding Sites Biochemical Biological Assay C57BL/6 Mouse Cattle Cell Culture Techniques Cell membrane Cells Characteristics Charge Circular Dichroism Spectroscopy Collaborations Data Dimerization EMSA Endothelial Cells Enzymes Epithelial Fatty Acids Fluorescence Fluorescence Resonance Energy Transfer Generations Genes Genetic Polymorphism Grant Homo Human Cloning Hydrogen Peroxide Hydroxyl Radical Hyperoxia In Vitro Injury Knock-in Mouse Knock-out Knockout Mice Lipid Peroxides Liposomes Lung MCF7 cell Measurement Membrane Messenger RNA Metabolism Methods Modeling Molecular Mus Mutagenesis Mutate Mutation Oxidants Oxidative Stress Paraquat Peroxidases Phospholipase Phospholipase A2 Phospholipids Play Promoter Regions Property Proteins Rat Protein Reaction Reactive Oxygen Species Recombinant Proteins Regulation Resistance Response Elements Role Selenium Site Site-Directed Mutagenesis Stress Structure Surface Surface Plasmon Resonance System Systems Analysis Testing Tryptophan ascorbate base deletion analysis enzyme activity enzyme structure glutathione peroxidase mouse model mutant novel overexpression oxidant stress peroxiredoxin phosphatidylcholine hydroperoxide repaired response surfactant transcription factor

项目摘要

This project will evaluate the properties of a novel glutathione peroxidase enzyme that is considerably enriched in lungs. This recently described protein (called 1-cys Peroxiredoxin or Peroxiredoxin 6, Prdx 6) represents the only non-selenium glutathione peroxidase of relatively high specific activity. An important characteristic in the spectrum activity for this enzyme is its ability, unlike classical glutathione peroxidase, to reduce phospholipid hydroperoxides. During the present grant Support as an R-01, we have demonstrated induction of Prdx 6 during hyperoxic stress and provided evidence that the enzyme protects against oxidant stress in cell culture and whole animal studies. Further, we have developed methods for isolation of Prdx 6 protein from rat and bovine lungs and have established expression systems for generation of active recombinant protein. We postulate that reduction of oxidized membrane phospholipids represents the basis for the anti-oxidant effect of the enzyme. Specific Aim I will utilize mouse models of altered Prdx 6 expression to continue our studies of the role of Prdx 6 in anti-oxidant defense. One of the models, Prdx 6 knock-out mice, was developed during the present period of grant support. The second Specific Aim will investigate control of Prdx 6 expression with special emphasis on the anti-oxidant response element and Nrf 2. Specific Aim 3 will evaluate substrate-protein interactions with a special focus on phospholipid binding by the protein as a requirement for the phospholipid hydroperoxide peroxidase activity. These studies will utilize site directed mutagenesis based on deduction from the crystal structure of the enzyme. Specific Aim 4 will utilize cellular systems for analysis of the role of nGST in activation of Prdx 6 activity. The proposed studies will provide a coordinated effort to investigate the role of this novel anti-oxidant enzyme in lung defense against oxidant stress and will provide new information concerning the biochemical regulation of its enzymatic activity.

该项目将评估一种新型的谷胱甘肽过氧化物酶的特性，该酶在肺中大大富集。这是最近描述的蛋白质（称为1-CYS过氧蛋白或过氧蛋白6，PRDX 6）代表了相对较高特异性活性的唯一非谷胱甘肽过氧化物酶。这种酶的光谱活性中的一个重要特征是它的能力，与经典的谷胱甘肽过氧化物酶不同，可以减少磷脂氢过氧化物。在当前的R-01授予支持期间，我们在高氧气应激期间证明了PRDX 6的诱导，并提供了证据表明该酶可以防止细胞培养和整个动物研究中的氧化应激。此外，我们开发了从大鼠和牛肺中分离PrDX 6蛋白的方法，并建立了用于生成活性的表达系统重组蛋白。我们假设氧化膜磷脂的还原是该酶抗氧化作用的基础。具体目的我将利用改变PRDX 6表达的小鼠模型继续我们对PRDX 6在抗氧化剂防御中的作用的研究。其中一种模型是PRDX 6敲除小鼠，在目前的赠款支持期间开发了。第二个具体目的将研究对PRDX 6表达的控制，并特别强调抗氧化剂反应元件和NRF 2。具体目标3将评估底物 - 蛋白质相互作用，特别关注蛋白质的磷脂结合，作为对磷脂脂脂氧化氧化物过氧化物过氧化物酶活性的要求。这些研究会利用基于从酶的晶体结构中扣除的位置诱变。具体目标4将利用细胞系统来分析NGST在激活PRDX 6活性中的作用。拟议的研究将提供协调的努力，以研究这种新型抗氧化酶在肺防御抗氧化应激中的作用，并将提供有关其酶活性生化调节的新信息。