Department of Pathology, Faculty of Medical Sciences, University of Campinas (UNICAMP), Rua Tessália Vieira de Camargo, 126, 13083-887, Campinas, SP, Brazil.
Department of Pathology, Faculty of Medical Sciences, University of Campinas (UNICAMP), Rua Tessália Vieira de Camargo, 126, 13083-887, Campinas, SP, Brazil.
Arch Biochem Biophys. 2021 Aug 15;707:108934. doi: 10.1016/j.abb.2021.108934. Epub 2021 May 25.
HO is endogenously generated and its removal in the matrix of skeletal muscle mitochondria (SMM) is dependent on NADPH likely provided by NAD(P) transhydrogenase (NNT) and isocitrate dehydrogenase (IDH2). Importantly, NNT activity is linked to mitochondrial protonmotive force. Here, we demonstrate the presence of NNT function in detergent-solubilized and intact functional SMM isolated from rats and wild type (Nnt) mice, but not in SMM from congenic mice carrying a mutated NNT gene (Nnt). Further comparisons between SMM from both Nnt mouse genotypes revealed that the NADPH supplied by NNT supports up to 600 pmol/mg/min of HO removal under selected conditions. Surprisingly, SMM from Nnt mice removed exogenous HO at wild-type levels and exhibited a maintained or even decreased net emission of endogenous HO when substrates that support Krebs cycle reactions were present (e.g., pyruvate plus malate or palmitoylcarnitine plus malate). These results may be explained by a compensation for the lack of NNT, since the total activities of concurrent NADP-reducing enzymes (IDH2, malic enzymes and glutamate dehydrogenase) were ~70% elevated in Nnt mice. Importantly, respiratory rates were similar between SMM from both Nnt genotypes despite differing NNT contributions to HO removal and their implications for an evolving concept in the literature are discussed. We concluded that NNT is capable of meaningfully sustaining NADPH-dependent HO removal in intact SMM. Nonetheless, if the available substrates favor non-NNT sources of NADPH, the HO removal by SMM is maintained in Nnt mice SMM.
HO 是内源性产生的，其在骨骼肌线粒体基质（SMM）中的去除依赖于 NADPH，可能由 NAD(P)转氢酶（NNT）和异柠檬酸脱氢酶（IDH2）提供。重要的是，NNT 活性与线粒体质子动力有关。在这里，我们证明了在从大鼠和野生型（Nnt）小鼠中分离的去污剂可溶和完整功能的 SMM 中存在 NNT 功能，但在携带突变 NNT 基因的同基因小鼠（Nnt）的 SMM 中不存在。对来自两种 Nnt 基因型的 SMM 之间的进一步比较表明，NNT 提供的 NADPH 在选定条件下支持高达 600 pmol/mg/min 的 HO 去除。令人惊讶的是，Nnt 小鼠的 SMM 以野生型水平去除外源性 HO，并在存在支持三羧酸循环反应的底物（例如，丙酮酸加苹果酸或棕榈酰肉碱加苹果酸）时表现出维持或甚至降低内源性 HO 的净发射。这些结果可以通过对 NNT 缺乏的补偿来解释，因为同时 NADP 还原酶（IDH2、苹果酸酶和谷氨酸脱氢酶）的总活性在 Nnt 小鼠中升高了约 70%。重要的是，尽管两种 Nnt 基因型的 SMM 的 NNT 对 HO 去除的贡献不同，但其对文献中不断发展的概念的影响不同，但它们的呼吸率相似。我们得出结论，NNT 能够在完整的 SMM 中有效地维持 NADPH 依赖性的 HO 去除。尽管如此，如果可用的底物有利于非 NNT 来源的 NADPH，则 Nnt 小鼠的 SMM 中的 HO 去除仍能维持。
