DETECT, ID & CHARACTERIZE BENZENE ADDUCTED MOUSE LIVER & BONE MARROW PROTEINS

检测、识别

• 批准号: 6308826
• 负责人: ALMA L BURLINGAME
• 金额: $ 0.99万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-01 至 2002-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6308826
• 关键词: Mammalia; biological products; biomedical resource; drug adverse effect; environmental toxicology; proteins; spectrometry; structural biology; technology /technique

By combining the ability of accelerator mass spectrometry to detect low levels of 14C with the high sensitivity of high energy collision-induced dissociation analysis for peptide sequence determination, we have developed a novel approach to detect and characterize proteins withadducted xenobiotic compounds. In the present study, we are trying to identify benzene-protein adducts purified from mouse liver and bone marrow cells. Benzene is a commonly occurring toxic compound and is present in combustion products, gasoline and tobacco smoke. It is known that chronic exposure to benzene leads to degeneration of the bone marrow and leukemia. Further, it has also been shown that benzene-protein adducts are formed via metabolism of benzene. Bioactivation of benzene occurs in the liver through oxidation by P450to phenol. However, several other reactive metabolites are formed as well, such as benzoquinones, catechol hydroquinone and muconaldehyde. The hematotoxic effect of benzene has been attributed to these benzene intermediates rather than benzene or phenol. The aim of this workis to characterize the protein adducts formed with benzene and to elucidate their possible importance for the development of leukemia and other benzene-induced pathological conditions. Crude liver homogenates obtained from mice injected with low amounts (500 ng/kg body weight) of 14C-benzene are fractionated by SDS-PAGE and the 14C-enriched proteins are detected by accelerator MS. The proteins of interest are subjected to in-gel trypsin digestion and the extracted peptides sequenced by high energy CID analysis.

通过结合加速器质谱分析的能力， 以高能量的高灵敏度检测低水平的14 C 肽序列碰撞诱导解离分析 决心，我们已经开发出一种新的方法来检测和 用加合的异生物质化合物表征蛋白质。 在 目前的研究，我们正试图确定苯-蛋白质加合物 从小鼠肝脏和骨髓细胞中纯化。 苯是一种 常见的有毒化合物，存在于燃烧中 产品、汽油和烟草烟雾。 据了解，慢性 接触苯会导致骨髓退化， 白血病 此外，还表明苯-蛋白质 加合物通过苯的代谢形成。 生物激活 苯在肝脏中通过P450氧化成苯酚而发生。 然而，也形成了几种其他反应性代谢物，例如 如苯醌、邻苯二酚对苯二酚和粘康醛。 的 苯血液毒性作用归因于这些苯 中间体而不是苯或苯酚。 这项工作的目的是 表征与苯形成的蛋白质加合物， 阐明它们对白血病发展的可能重要性 和其他苯引起的病理状况。 粗肝 从注射少量（500 ng/kg）的小鼠获得的匀浆 用SDS-PAGE分离14 C-苯， 通过加速器MS检测14 C富集的蛋白质。 对感兴趣的样品进行凝胶内胰蛋白酶消化， 通过高能CID分析测序的肽。