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

通过结合加速器质谱学的能力 利用高能的高灵敏度检测低水平的14C 多肽序列的碰撞诱导解离分析 决心，我们开发了一种新的方法来检测和 用加成的外源化合物表征蛋白质。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提纯。苯是一种 常见的有毒化合物，存在于燃烧过程中 产品、汽油和烟草烟雾。众所周知，慢性病 暴露在苯中会导致骨髓退化和 白血病。此外，研究还表明，苯蛋白 加合物是通过苯的代谢形成的。生物活化法 苯在肝脏中被P450氧化成苯酚。 然而，也会形成其他几种反应性代谢物，如 苯二酚、邻苯二酚对苯二酚和邻苯二醛。这个 苯的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 对与苯形成的蛋白质加合物进行表征 阐明它们在白血病发生发展中可能的重要作用 以及其他由苯引起的病理情况。粗肝 小鼠注射低剂量(500 ng/kg)获得的匀浆 体重)的14C-苯用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳法进行分级 用加速器MS检测富含14C的蛋白质。 感兴趣的是胶内胰酶消化和提取的 多肽的高能CID分析测序。

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.