Human dihydrolipoamide dehydrogenase (E3) deficiency: Novel insights into the structural basis and molecular pathomechanism.

This review summarizes our present view on the molecular pathogenesis of human (h) E3-deficiency caused by a variety of genetic alterations with a special emphasis on the moonlighting biochemical phenomena related to the affected (dihydro)lipoamide dehydrogenase (LADH, E3, gene: dld), in particular the generation of reactive oxygen species (ROS). E3-deficiency is a rare autosomal recessive genetic disorder frequently presenting with a neonatal onset and premature death; the highest carrier rate of a single pathogenic dld mutation (1:94-1:110) was found among Ashkenazi Jews. Patients usually die during acute episodes that generally involve severe metabolic decompensation and lactic acidosis leading to neurological, cardiological, and/or hepatological manifestations. The disease owes its severity to the fact that LADH is the common E3 subunit of the alpha-ketoglutarate (KGDHc), pyruvate (PDHc), and branched-chain α-keto acid dehydrogenase complexes and is also part of the glycine cleavage system, hence the malfunctioning of LADH simultaneously incapacitates several central metabolic pathways. Nevertheless, the clinical pictures are usually not unequivocally portrayed through the loss of LADH activities and imply auxiliary mechanisms that exacerbate the symptoms and outcomes of this disorder. Enhanced ROS generation by disease-causing hE3 variants as well as by the E1-E2 subcomplex of the hKGDHc likely contributes to selected pathogeneses of E3-deficiency, which could be targeted by specific drugs or antioxidants; lipoic acid was demonstrated to be a potent inhibitor of ROS generation by hE3 in vitro. Flavin supplementation might prove to be beneficial for those mutations triggering FAD loss in the hE3 component. Selected pathogenic hE3 variants lose their affinity for the E2 component of the hPDHc, a mechanism which warrants scrutiny also for other E3-haboring complexes.

Complex contribution of cyclophilin D to Ca2+-induced permeability transition in brain mitochondria, with relation to the bioenergetic state.

Cyclophilin D (cypD)-deficient mice exhibit resistance to focal cerebral ischemia and to necrotic but not apoptotic stimuli. To address this disparity, we investigated isolated brain and in situ neuronal and astrocytic mitochondria from cypD-deficient and wild-type mice. Isolated mitochondria were challenged by high Ca(2+), and the effects of substrates and respiratory chain inhibitors were evaluated on permeability transition pore opening by light scatter. In situ neuronal and astrocytic mitochondria were visualized by mito-DsRed2 targeting and challenged by calcimycin, and the effects of glucose, NaCN, and an uncoupler were evaluated by measuring mitochondrial volume. In isolated mitochondria, Ca(2+) caused a large cypD-dependent change in light scatter in the absence of substrates that was insensitive to Ruthenium red or Ru360. Uniporter inhibitors only partially affected the entry of free Ca(2+) in the matrix. Inhibition of complex III/IV negated the effect of substrates, but inhibition of complex I was protective. Mitochondria within neurons and astrocytes exhibited cypD-independent swelling that was dramatically hastened when NaCN and 2-deoxyglucose were present in a glucose-free medium during calcimycin treatment. In the presence of an uncoupler, cypD-deficient astrocytic mitochondria performed better than wild-type mitochondria, whereas the opposite was observed in neurons. Neuronal mitochondria were examined further during glutamate-induced delayed Ca(2+) deregulation. CypD-knock-out mitochondria exhibited an absence or a delay in the onset of mitochondrial swelling after glutamate application. Apparently, some conditions involving deenergization render cypD an important modulator of PTP in the brain. These findings could explain why absence of cypD protects against necrotic (deenergized mitochondria), but not apoptotic (energized mitochondria) stimuli.