Reverse and Forward Electron Flow-Induced H2O2 Formation Is Decreased in α-Ketoglutarate Dehydrogenase (α-KGDH) Subunit (E2 or E3) Heterozygote Knock Out Animals

Abstract

α-ketoglutarate dehydrogenase complex (KGDHc), or 2-oxoglutarate dehydrogenase complex (OGDHc) is a rate-limiting enzyme in the tricarboxylic acid cycle, that has been identified in neurodegenerative diseases such as in Alzheimer’s disease. The aim of the present study was to establish the role of the KGDHc and its subunits in the bioenergetics and reactive oxygen species (ROS) homeostasis of brain mitochondria. To study the bioenergetic profile of KGDHc, genetically modified mouse strains were used having a heterozygous knock out (KO) either in the dihydrolipoyl succinyltransferase (DLST+/−) or in the dihydrolipoyl dehydrogenase (DLD+/−) subunit. Mitochondrial oxygen consumption, hydrogen peroxide (H2O2) production, and expression of antioxidant enzymes were measured in isolated mouse brain mitochondria. Here, we demonstrate that the ADP-stimulated respiration of mitochondria was partially arrested in the transgenic animals when utilizing α-ketoglutarate (α-KG or 2-OG) as a fuel substrate. Succinate and α-glycerophosphate (α-GP), however, did not show this effect. The H2O2 production in mitochondria energized with α-KG was decreased after inhibiting the adenine nucleotide translocase and Complex I (CI) in the transgenic strains compared to the controls. Similarly, the reverse electron transfer (RET)-evoked H2O2 formation supported by succinate or α-GP were inhibited in mitochondria isolated from the transgenic animals. The decrease of RET-evoked ROS production by DLST+/− or DLD+/− KO-s puts the emphasis of the KGDHc in the pathomechanism of ischemia-reperfusion evoked oxidative stress. Supporting this notion, expression of the antioxidant enzyme glutathione peroxidase was also decreased in the KGDHc transgenic animals suggesting the attenuation of ROS-producing characteristics of KGDHc. These findings confirm the contribution of the KGDHc to the mitochondrial ROS production and in the pathomechanism of ischemia-reperfusion injury.

The emerging importance of the α-keto acid dehydrogenase complexes in serving as intracellular and intercellular signaling platforms for the regulation of metabolism

“…Studies that used permeabilized rat skeletal muscle mitochondria, intact and permeabilized mouse liver mitochondria, cultured macrophages, and heart mitochondria has revealed PDHc, KGDHc, BCKDHc, and OADHc display rates of mtO 2 •- /mtH 2 O 2 greater than complex I [ 1 , 49 , 83 , 84 , [86] , [87] , [88] , [89] ]. Horvath et al reported KGDHc is also an important mtO 2 •- /mtH 2 O 2 generator in mouse brain tissue when the ETC is operating under RET conditions [ 90 ]. In this case, mitochondrial fuels like succinate and glycerol-3-phosphate, which by-pass the Krebs cycle and donate electrons directly to coenzyme Q 10 (CoQ) in the ETC, generate mtO 2 •- /mtH 2 O 2 by KGDHc by producing NADH at complex I [ 90 ].…”

“…Horvath et al reported KGDHc is also an important mtO 2 •- /mtH 2 O 2 generator in mouse brain tissue when the ETC is operating under RET conditions [ 90 ]. In this case, mitochondrial fuels like succinate and glycerol-3-phosphate, which by-pass the Krebs cycle and donate electrons directly to coenzyme Q 10 (CoQ) in the ETC, generate mtO 2 •- /mtH 2 O 2 by KGDHc by producing NADH at complex I [ 90 ]. Together, a significant amount of evidence has been generated demonstrating the KDHc family of enzymes are main mtO 2 •- /mtH 2 O 2 sources in mammalian mitochondria.…”

“…The observation that NADH accumulation can augment mtO 2 •- /mtH 2 O 2 formation by KGDHc was demonstrated by Starkov et al using rotenone, a complex I inhibitor [ 76 ]. Similarly, Horvath et al successfully showed KGDHc can produce large quantities of mtO 2 •- /mtH 2 O 2 when NADH is formed by the reversal of complex I, which can be prevented with rotenone [ 90 ]. Similar findings were made by Quinlan et al where addition of rotenone significantly augmented mtO 2 •- /mtH 2 O 2 production in rat muscle mitochondria oxidizing pyruvate or BCKAs [ 87 ].…”

Fatty acid oxidation drives hydrogen peroxide production by α-ketoglutarate dehydrogenase

“…Further, complex I and III of the ETC are often viewed as the main sources of mH 2 O 2 , but this convention has been challenged by the growing evidence showing PDH and KGDH are important mH 2 O 2 generators. Several studies demonstrated KGDH has a high mH 2 O 2 production rate in synaptosomes and isolated mitochondria (Horvath et al , 2022; Starkov et al , 2004; Tretter & Adam-Vizi, 2004). It was reported later that KGDH is 8x more effective at mH 2 O 2 generation when compared to complex I in muscle (Quinlan et al , 2014).…”

Oxidative Stress, Neuroinflammation and Neurodegeneration: The Chicken, the Egg and the Dinosaur

“…Moreover, it was demonstrated here that the redox-sensitive protein DJ-1 is required to protect the retina and retinal pigment epithelium (RPE) from oxidative-stress-induced degeneration [ 16 ]. This Special Issue also gathered studies focused on the identification of new biomarkers for AD preclinical diagnosis [ 17 ], of new treatments based on the administration of hydroxocobalamin (Hb, vitamin B 12 analog) to prevent cytoplasmic aggregation of TDP-43 observed in many neurodegenerative diseases [ 18 ], of a new mechanism linking astrocyte-derived oxidative stress to motor-neuron damage in ALS [ 19 ], and on the role of the alpha–ketoglutarate dehydrogenase complex (KGDHC), a rate-limiting enzyme in the tricarboxylic acid cycle, whose activity is strikingly reduced in AD [ 16 ], in the bioenergetics and reactive oxygen species (ROS) homeostasis of brain mitochondria [ 20 ].…”