BIOCHEMISTRY AND HEPATIC TOXICITY OF CARBON RADICALS

描述（由申请人提供）：自由基过程涉及越来越多的病理，包括主要的健康问题，如癌症和动脉粥样硬化。然而，尽管在过去几年中取得了令人印象深刻的进展，但对蛋白质自由基的基本生物化学仍然不完全了解。工作提出了应用基于关键进步期到期的支持,包括(a)识别的残留在乳过氧化物酶催化血红素组的共价结合,(b)识别特定的残留在乳过氧化物酶转化成自由基中心与H202经酶反应,(c)突变的辣根过氧化物酶酶,模仿能力的哺乳动物氧化酵素其血红素共价结合,(d)在肌红蛋白系统中证明酪氨酸氧化为自由基取决于与氧化中心距离以外的其他因素。目前的项目主要集中在蛋白质自由基生物化学的两个方面：(a)羧酸自由基（一种高度被忽视的物种）在过氧化物酶功能中的可能形成和作用，以及(b)不寻常的分子内交联的形成机制及其功能后果。本项目的第一个目标是通过研究辣根过氧化物酶模型系统来确定羧酸基团在血红素-蛋白交联反应中的作用。第二个目的是使用相同的模型来阐明自由基在髓过氧化物酶蛋氨酸-乙烯基键形成中的作用。第三个目的是澄清为什么共价血红素结合对哺乳动物过氧化物酶功能很重要。第四个目的是研究活性位点羧酸基团的作用和过氧化氢酶-过氧化物酶中不寻常的交联形成的机制。最后的目标是采用肌红蛋白模型系统和附加的酚来探索确定蛋白质中哪些酪氨酸被氧化的关系。这些研究将揭示生理和病理过程中过氧化碳自由基的产生和命运，并为调节或抑制这些过程提供新的思路。

DESCRIPTION (provided by applicant): Free radical processes have been implicated in a growing range of pathologies, including major health problems such as cancer and atherosclerosis. Nevertheless, despite impressive progress in the past few years, the basic biochemistry of protein radicals continues to be incompletely understood. The work proposed in this application rests on key advances made in the expiring period of support, including (a) identification of the residues in autocatalytic covalent binding of the heme group in lactoperoxidase, (b) Identification of specific residues in lactoperoxidase that are converted to free radicals centers upon reaction of the enzyme with H202, (c) mutation of horseradish peroxidase into an enzyme that mimics the mammalian peroxidases in its ability to bind its heme covalently, and (d) demonstration in a metmyoglobin system that the oxidation of tyrosines to radicals depends on factors other than distance from the oxidizing center. The present project focuses on two aspects of protein radical biochemistry: (a) the possible formation and role of carboxylate radicals, a highly neglected species, in peroxidase function, and (b) the mechanism of formation of unusual intramolecular cross-links and their functional consequences. The first aim of the present project is to define the role of the carboxylate group in the heme-protein cross-linking reaction through studies of the horseradish peroxidase model system. The second aim is to use the same model to clarify the role of radicals in formation of the methionine-vinyl bond of myeloperoxidase. The third aim is to clarify why covalent heme binding is important for mammalian peroxidase function. The fourth aim is to examine the role of the active site carboxylate group and the mechanism(s) by which the unusual cross-links in catalase-peroxidase enzymes are formed. A final goal is to employ a metmyoglobin model system with appended phenols to explore the relationships which determine which tyrosines are oxidized in a protein. These studies should shed light on the generation and fates of peroxidatively generated carbon radicals in physiological and pathological processes and provide insights into approaches for the modulation or suppression of such processes.