REDOX INTERACTIONS OF DT DIAPHORASE AND VITAMIN E

NAD(P)H：醌氧化还原酶（NQO1, EC 1.699.2, DT-diaphorase）是一种普遍存在的酶，可对多种醌底物进行2电子还原，并在抗氧化反应中被诱导。尽管NQO1的“生理”底物尚不清楚，但我们最近发现NQO1可以将内源性α -生育酚（α -维生素E）氧化产物α -生育酚醌(α -生育酚对苯二酚（α -生育酚对苯二酚）还原为α -生育酚对苯二酚（α -生育酚对苯二酚），而α -生育酚对苯二酚可以从α -生育酚自由基中再生α -生育酚。α th再生系统被认为是细胞抗氧化保护的重要贡献者。我们假设NQO1的一个基本功能是为α的氧化还原循环提供还原等价物，从而维持细胞抗氧化保护，防止细胞膜氧化损伤的传播。这种作用可能涉及nq1本身或TQH2对单电子氧化生育酚自由基或2电子氧化生育酚自由基的还原。为了验证这些假设，我们提出：目的1)表征TQH2和NQO1在单电子α氧化还原循环中的参与。将在脂质体和膜系统中研究直接由NQO1或通过TQH2将生育酚自由基还原为α。目的2)表征TQH2和NQO1在双电子α氧化还原循环中的参与。将在脂质体和膜模型中研究直接或通过TQH2还原8a-取代生育激素为α。目的3)表征NQO1对完整细胞中α氧化还原状态和抗氧化转换的影响。这些研究将使用CHO细胞和表达NQO1多态性的人类细胞系。α及其氧化产物的水平和分布将作为氧化应激细胞中NQO1活性的函数来测量。这些研究将阐明NQO1的内源性抗氧化作用，并确定自然界最普遍的膜抗氧化剂α的抗氧化化学如何与细胞代谢氧化还原平衡联系在一起。这些问题的答案对于提高我们对细胞抗氧化防御及其在人类健康和疾病中的作用之间相互作用的理解至关重要。

NAD(P)H:quinone oxidoreductase (NQO1, EC 1.699.2, DT-diaphorase) is a ubiquitous enzyme that carries out 2-electron reduction of numerous quinone substrates and is induced in antioxidant responses. Although a "physiologic" substrate for NQO1 had been obscure, we observed recently that NQO1 reduces the endogenous alpha-tocopherol (alphaTH, vitamin E) oxidation product alpha- tocopherolquinone (TQ) to alpha-tocopherolhydroquinone (TQH2), which can, in turn, regenerate alphaTH from the alpha- tocopheroxyl radical. An alphTH regenerating system is thought to be an essential contributor to cellular antioxidant protection. We hypothesize that an essential function of NQO1 is to supply reducing equivalents to redox cycles of alphaTh, thus sustaining cellular antioxidant protection against propagation of oxidative damage in membranes. This action may involve either reduction of one-electron oxidized tocopheroxyl radicals or 2- electron oxidized tocopherones either by NQO1 itself or by TQH2. To test these hypotheses, we propose: Aim 1) To characterize the participation of TQH2 and NQO1 in a one-electron alphaTH redox cycle. Reduction of the tocopheroxyl radical to alphaTH either directly by NQO1 or via TQH2 will be studied in liposome and membrane systems. Aim 2) To characterize the participation of TQH2 and NQO1 in a two-electron alphaTh redox cycle. Reduction of 8a-substituted tocopherones to alphaTH either directly or via TQH2 will be studied in liposome and membrane models. Aim 3) To characterize the impact of NQO1 on alphaTh redox status and antioxidant turnover in intact cells. These studies will employ CHO cells and human cell lines expressing NQO1 polymorphisms. The levels and distribution of alphaTH and its oxidation products will be measured as a function of NQO1 activity in cells subjected to oxidative stress. These studies will clarify the endogenous antioxidant role of NQO1 and determine how the antioxidant chemistry of alphaTH, Nature's most ubiquitous membrane antioxidant, is tied to cellular metabolic redox balance. Answers to these questions are essential to improving our understanding of the interplay between cellular antioxidant defenses and their roles in human health and disease.