NAD(P)H quinone oxidoreductase 1 (NQO1)-mediated bypass of mitochondrial electron transport chain with artificial and endogenous substrates

摘要 一系列罕见和常见的疾病与线粒体功能障碍和相关的氧化还原有关 不平衡。通过降低细胞内NADH/NAD+比率可以恢复潜在的氧化还原失衡 被视为在多种疾病状态下的一种极其有用的推广策略。NAD(P)H：苯二酚 氧化还原酶1(Oxoreductase 1，NQO1)是一种可溶性的细胞质酶，主要被认为是一种异源生物. 代谢酶，或抗癌药物的生物激活剂，以降低NAD(P)H的当量为代价。 有趣的是，一些NQO1人工底物，主要是萘醌，当还原时能够 随后将它们的电子捐献给络合物I下游的线粒体电子传输链。 因此，这种NQO1介导的替代电子转移是一种有吸引力的缓解还原应力的策略 并支持ATP动态平衡，因为它依赖于内源酶，并且只需要添加各自的 萘醌类化合物。然而，能够被NQO1还原的萘醌是天然产物或 合成氧化还原支架(如艾地苯酮)，我们目前缺乏关于NQO1内源底物的信息 以及它在细胞氧化还原代谢中的地位。为了弥合这一知识鸿沟，我们将使用基于活动的新陈代谢 用重组NQO1分析以鉴定与此相互转化的细胞内源性代谢物 酵素。接下来，我们将重建NQO1介导的电子转移与不同的萘醌在 分离线粒体，并将研究这个非标准的还原当量入口点的生物能量学。 这将使我们能够严格地表征萘醌和相关的氧化还原活性分子的能力。 在不引发氧化应激的情况下，安全地绕过受损的线粒体电子传输链。我们目前的情况 这种方法将第一次允许我们识别生理NQO1底物，并帮助我们更好地重建 NQO1介导的电子传递。这项工作最终将为开发治疗药物铺平道路。 基于氧化还原活性的小分子可以缓解还原压力的模式。

Abstract A wide range of rare and common diseases are linked to mitochondrial dysfunction and associated redox imbalance. Restoration of the underlying redox imbalance by decreasing the cellular NADH/NAD+ ratio could be seen as an extremely useful generalizable strategy in the context of multiple disease states. NAD(P)H:quinone oxidoreductase 1 (NQO1) is a soluble cytoplasmic enzyme that has been mostly viewed as a xenobiotic- metabolizing enzyme, or a bioactivator of cancer drugs at the expense of reducing equivalents of NAD(P)H. Interestingly, some of NQO1 artificial substrates, mostly naphthoquinones, when reduced are capable of subsequently donating their electrons to the mitochondrial electron transport chain downstream of Complex I. This NQO1-mediated alternative electron transfer is therefore an attractive strategy to alleviate reductive stress and support ATP homeostasis as it depends on an endogenous enzyme and only requires addition of respective naphthoquinones. However, naphthoquinones capable of being reduced by NQO1 are either natural products or synthetic redox scaffolds (e.g. idebenone), and we currently lack information on endogenous substrates of NQO1 and its place in cellular redox metabolism. To close this knowledge gap, we will use activity-based metabolomic profiling with recombinant NQO1 to identify cellular endogenous metabolites that are interconverted by this enzyme. Next, we will reconstitute the NQO1-mediated electron transfer with various naphthoquinones in isolated mitochondria and will study the bioenergetics of this non-canonical point of entry of reducing equivalents. This will allow us to rigorously characterize naphthoquinones and related redox-active molecules for their ability to safely bypass a corrupted mitochondrial electron transport chain without inducing oxidative stress. Our current approach will, for the first time, allow us to identify physiological NQO1 substrates and help us better reconstruct the NQO1-mediated electron transfer. This work will ultimately pave the way for developing therapeutic modalities that are based on redox-active small molecules that can alleviate reductive stress.