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

NAD(P)H 醌氧化还原酶 1 (NQO1) 介导的人工和内源底物线粒体电子传递链旁路

• 批准号: 10789749
• 负责人: Valentin Cracan
• 金额: $ 52.97万
• 依托单位: SCINTILLON INSTITUTE FOR PHOTOBIOLOGY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-30 至 2025-09-29
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10789749
• 关键词: Address; Analytical Chemistry; Antineoplastic Agents; Biochemical; Biochemical Pathway; Bioenergetics; Biological Assay; Biology; Bypass; Cells; Coenzymes; Complement; Complex; Consumption; Cytoplasm; Disease; Electron Transport; Electron Transport Complex III; Electrons; Energy Metabolism; Enzymes; Equilibrium; Genes; Goals; Homeostasis; Human; Hydrogen Peroxide; Incubated; Individual; Investigation; Knowledge; Lesion; Link; Mammalian Cell; Mass Spectrum Analysis; Mediating; Membrane Potentials; Metabolic; Metabolism; Mitochondria; Mitochondrial Diseases; Modality; Molecular; Monitor; Mus; Mutation; NADH; NADP; NQO1 gene; Naphthoquinones; Natural Products; Natural regeneration; Oxidation-Reduction; Oxidative Phosphorylation; Oxidative Stress Induction; Oxidoreductase; Oxygen; Pathology; Physiological; Physiology; Production; Proteins; Quinones; Rare Diseases; Reaction; Recombinants; Research; Role; Side; Source; Stress; Superoxides; Suspensions; System; Testing; Therapeutic; Therapeutic Intervention; Time; Tissues; Ubiquinone; Variant; Work; Xenobiotics; antioxidant enzyme; cancer therapy; candidate validation; combat; design; dimer; experimental study; idebenone; in vitro Assay; inhibitor; metabolomics; mitochondrial dysfunction; mitochondrial membrane; novel; overexpression; reconstitution; restoration; scaffold; small molecule

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.

摘要 一系列罕见和常见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+比值， 在多种疾病状态的背景下，这被视为一种非常有用的可推广策略。NAD（P）H：醌 氧化还原酶1（NQO 1）是一种可溶性细胞质酶，主要被视为异生素- 代谢酶，或癌症药物的生物活化剂，以减少NAD（P）H的当量为代价。 有趣的是，一些NQO 1人工底物，主要是萘醌，当还原时能够 随后将它们的电子贡献给复合物I下游的线粒体电子传递链。 因此，这种NQO 1介导的交替电子转移是减轻还原应激的有吸引力的策略 并支持ATP稳态，因为它依赖于内源性酶，仅需要添加相应的 萘醌。然而，能够被NQO 1还原的萘醌是天然产物， 合成氧化还原支架（例如艾地苯醌），我们目前缺乏关于NQO 1内源性底物的信息 以及它在细胞氧化还原代谢中的地位。为了缩小这一知识差距，我们将使用基于活性的代谢组学 用重组NQO 1进行谱分析，以鉴定通过这种方法相互转化的细胞内源性代谢物。 酵素接下来，我们将用不同的萘醌重建NQO 1介导的电子转移， 分离的线粒体，并将研究这个非规范的还原当量进入点的生物能量学。 这将使我们能够严格表征萘醌和相关的氧化还原活性分子的能力， 安全地绕过损坏的线粒体电子传递链而不诱导氧化应激。我们目前 这种方法将首次使我们能够识别生理NQO 1底物，并帮助我们更好地重建 NQO 1介导的电子转移。这项工作最终将为开发治疗性药物铺平道路。 基于可以减轻还原性应激的氧化还原活性小分子的模式。