PROTEIN HYDROPEROXIDES AS SECONDARY OXIDANTS IN HYDROXYL RADICAL FOOTPRINTING

蛋白质氢过氧化物作为羟基自由基足迹中的二级氧化剂

• 批准号: 8168868
• 负责人: JOSHUA S SHARP
• 金额: $ 3.37万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-10 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8168868
• 关键词: Amides; Amino Acids; Computer Retrieval of Information on Scientific Projects Database; Electrons; Funding; Grant; Hydrogen Peroxide; Hydroxyl Radical; Institution; Leucine; Liquid Chromatography; Mass Spectrum Analysis; Methionine; Methods; Oxidants; Peptides; Preparation; Proteins; Research; Research Personnel; Resources; Series; Side; Solutions; Source; Sulfur; United States National Institutes of Health; catalase; design; oxidation; research study

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Previous studies of oxidation of leucine-containing peptides revealed the formation of a peptidyl hydroperoxide on the leucine side chain. Peptidyl hydroperoxides have never before been observed in hydroxyl radical footprinting experiments. It was previously thought that they were too unstable to survive the protein preparation and mass spectrometry analysis; however, their discovery in the leucine-containing peptide by mass spectrometry indicates that they are sufficiently stable to persist in solution. If they do exist in solution, they may be capable of performing direct two-electron oxidation of sulfur-containing side chains, causing false positives in hydroxyl radical footprinting experiments. Additionally, standard methods used to remove hydrogen peroxide, including catalase treatment and liquid chromatography would be ineffective at removing peptidyl hydroperoxides. We designed and carried out a series of experiments to determine if peptidyl hydroperoxides are common products of oxidation of various amino acids, to determine if these peptidyl hydroperoxides are capable of two-electron oxidation of methionine residues, and to determine if catalase treatment or scavenging with methionine amide is effective at eliminating the peptidyl hydroperoxides.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 先前关于含亮氨酸肽的氧化的研究表明，在亮氨酸侧链上形成了肽基氢过氧化物。 在羟基自由基足迹实验中从未观察到肽基氢过氧化物。 以前认为它们太不稳定了，无法在蛋白质制备和质谱分析中幸存下来。但是，它们通过质谱法在含亮氨酸的肽中发现的发现表明它们足够稳定以持续溶液。 如果它们确实存在于溶液中，则它们可能能够对含硫的侧链进行直接的两电子氧化，从而在羟基自由基足迹实验中引起假阳性。 此外，用于去除过氧化氢的标准方法，包括过氧化氢酶处理和液相色谱法将无效去除肽基氢过氧化物。 我们设计并进行了一系列实验，以确定肽基氢过氧化物是否是各种氨基酸氧化的常见产物，以确定这些肽基氢氧化物是否能够对甲基氨酸残基的两种电子氧化，并确定用甲基酰胺进行甲基化酰胺的氧化酶处理或有效地消除甲基水素。