Dataset Information

The negative impact of ?-ketoglutarate dehydrogenase complex deficiency on matrix substrate-level phosphorylation.

ABSTRACT: A decline in ?-ketoglutarate dehydrogenase complex (KGDHC) activity has been associated with neurodegeneration. Provision of succinyl-CoA by KGDHC is essential for generation of matrix ATP (or GTP) by substrate-level phosphorylation catalyzed by succinyl-CoA ligase. Here, we demonstrate ATP consumption in respiration-impaired isolated and in situ neuronal somal mitochondria from transgenic mice with a deficiency of either dihydrolipoyl succinyltransferase (DLST) or dihydrolipoyl dehydrogenase (DLD) that exhibit a 20-48% decrease in KGDHC activity. Import of ATP into the mitochondrial matrix of transgenic mice was attributed to a shift in the reversal potential of the adenine nucleotide translocase toward more negative values due to diminished matrix substrate-level phosphorylation, which causes the translocase to reverse prematurely. Immunoreactivity of all three subunits of succinyl-CoA ligase and maximal enzymatic activity were unaffected in transgenic mice as compared to wild-type littermates. Therefore, decreased matrix substrate-level phosphorylation was due to diminished provision of succinyl-CoA. These results were corroborated further by the finding that mitochondria from wild-type mice respiring on substrates supporting substrate-level phosphorylation exhibited ~30% higher ADP-ATP exchange rates compared to those obtained from DLST(+/-) or DLD(+/-) littermates. We propose that KGDHC-associated pathologies are a consequence of the inability of respiration-impaired mitochondria to rely on "in-house" mitochondrial ATP reserves.

SUBMITTER: Kiss G

PROVIDER: S-EPMC3659355 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

The negative impact of α-ketoglutarate dehydrogenase complex deficiency on matrix substrate-level phosphorylation.

Kiss Gergely G Konrad Csaba C Doczi Judit J Starkov Anatoly A AA Kawamata Hibiki H Manfredi Giovanni G Zhang Steven F SF Gibson Gary E GE Beal M Flint MF Adam-Vizi Vera V Chinopoulos Christos C

FASEB journal : official publication of the Federation of American Societies for Experimental Biology 20130308 6

A decline in α-ketoglutarate dehydrogenase complex (KGDHC) activity has been associated with neurodegeneration. Provision of succinyl-CoA by KGDHC is essential for generation of matrix ATP (or GTP) by substrate-level phosphorylation catalyzed by succinyl-CoA ligase. Here, we demonstrate ATP consumption in respiration-impaired isolated and in situ neuronal somal mitochondria from transgenic mice with a deficiency of either dihydrolipoyl succinyltransferase (DLST) or dihydrolipoyl dehydrogenase (D ...[more]

PMID: 23475850

Similar Datasets

Project description:It has become appreciated over the last several years that protein phosphorylation within the cardiac mitochondrial matrix and respiratory complexes is extensive. Given the importance of oxidative phosphorylation and the balance of energy metabolism in the heart, the potential regulatory effect of these classical signaling events on mitochondrial function is of interest. However, the functional impact of protein phosphorylation and the kinase/phosphatase system responsible for it are relatively unknown. Exceptions include the well-characterized pyruvate dehydrogenase and branched chain ?-ketoacid dehydrogenase regulatory system. The first task of this review is to update the current status of protein phosphorylation detection primarily in the matrix and evaluate evidence linking these events with enzymatic function or protein processing. To manage the scope of this effort, we have focused on the pathways involved in energy metabolism. The high sensitivity of modern methods of detecting protein phosphorylation and the low specificity of many kinases suggests that detection of protein phosphorylation sites without information on the mole fraction of phosphorylation is difficult to interpret, especially in metabolic enzymes, and is likely irrelevant to function. However, several systems including protein translocation, adenine nucleotide translocase, cytochrome c, and complex IV protein phosphorylation have been well correlated with enzymatic function along with the classical dehydrogenase systems. The second task is to review the current understanding of the kinase/phosphatase system within the matrix. Though it is clear that protein phosphorylation occurs within the matrix, based on (32)P incorporation and quantitative mass spectrometry measures, the kinase/phosphatase system responsible for this process is ill-defined. An argument is presented that remnants of the much more labile bacterial protein phosphoryl transfer system may be present in the matrix and that the evaluation of this possibility will require the application of approaches developed for bacterial cell signaling to the mitochondria.

Project description:BACKGROUND: Synthesis and apoenzyme attachment of lipoic acid have emerged as a new complex metabolic pathway. Mutations in several genes involved in the lipoic acid de novo pathway have recently been described (i.e., LIAS, NFU1, BOLA3, IBA57), but no mutation was found so far in genes involved in the specific process of attachment of lipoic acid to apoenzymes pyruvate dehydrogenase (PDHc), ?-ketoglutarate dehydrogenase (?-KGDHc) and branched chain ?-keto acid dehydrogenase (BCKDHc) complexes. METHODS: Exome capture was performed in a boy who developed Leigh disease following a gastroenteritis and had combined PDH and ?-KGDH deficiency with a unique amino acid profile that partly ressembled E3 subunit (dihydrolipoamide dehydrogenase / DLD) deficiency. Functional studies on patient fibroblasts were performed. Lipoic acid administration was tested on the LIPT1 ortholog lip3 deletion strain yeast and on patient fibroblasts. RESULTS: Exome sequencing identified two heterozygous mutations (c.875C?>?G and c.535A?>?G) in the LIPT1 gene that encodes a mitochondrial lipoyltransferase which is thought to catalyze the attachment of lipoic acid on PDHc, ?-KGDHc, and BCKDHc. Anti-lipoic acid antibodies revealed absent expression of PDH E2, BCKDH E2 and ?-KGDH E2 subunits. Accordingly, the production of 14CO2 by patient fibroblasts after incubation with 14Cglucose, 14Cbutyrate or 14C3OHbutyrate was very low compared to controls. cDNA transfection experiments on patient fibroblasts rescued PDH and ?-KGDH activities and normalized the levels of pyruvate and 3OHbutyrate in cell supernatants. The yeast lip3 deletion strain showed improved growth on ethanol medium after lipoic acid supplementation and incubation of the patient fibroblasts with lipoic acid decreased lactate level in cell supernatants. CONCLUSION: We report here a putative case of impaired free or H protein-derived lipoic acid attachment due to LIPT1 mutations as a cause of PDH and ?-KGDH deficiencies. Our study calls for renewed efforts to understand the mechanisms of pathology of lipoic acid-related defects and their heterogeneous biochemical expression, in order to devise efficient diagnostic procedures and possible therapies.

Dataset Information

The negative impact of ?-ketoglutarate dehydrogenase complex deficiency on matrix substrate-level phosphorylation.

Publications

The negative impact of α-ketoglutarate dehydrogenase complex deficiency on matrix substrate-level phosphorylation.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure