Abstract
Human NADH CoQ oxidoreductase is composed of a total of 43 subunits and has been demonstrated to be a major site for the production of superoxide by mitochondria. Incubation of rat heart mitochondria with ATP resulted in the phosphorylation of two mitochondrial membrane proteins, one with a M(r) of 6 kDa consistent with the NDUFA1 (MWFE), and one at 18kDa consistent with either NDUFS4 (AQDQ) or NDUFB7 (B18). Phosphorylation of both subunits was enhanced by cAMP derivatives and protein kinase A (PKA) and was inhibited by PKA inhibitors (PKAi). When mitochondrial membranes were incubated with pyruvate dehydrogenase kinase, phosphorylation of an 18kDa protein but not a 6kDa protein was observed. NADH cytochrome c reductase activity was decreased and superoxide production rates with NADH as substrate were increased. On the other hand, with protein kinase A-driven phosphorylation, NADH cytochrome c reductase was increased and superoxide production decreased. Overall there was a 4-fold variation in electron transport rates observable at the extremes of these phosphorylation events. This suggests that electron flow through complex I and the production of oxygen free radicals can be regulated by phosphorylation events. In light of these observations we discuss a potential model for the dual regulation of complex I and the production of oxygen free radicals by both PKA and PDH kinase.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenosine Triphosphate / pharmacology
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Animals
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Blotting, Western
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Cyclic AMP / metabolism
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Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
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Cyclic AMP-Dependent Protein Kinases / pharmacology*
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Electron Transport Complex I
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Enzyme Activation
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Firefly Luciferin / metabolism
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Humans
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Imidazoles*
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Membrane Proteins / antagonists & inhibitors
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Membrane Proteins / metabolism
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Mitochondria, Heart / drug effects*
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Mitochondria, Heart / enzymology
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NAD / metabolism
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NADH Dehydrogenase / metabolism
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NADH, NADPH Oxidoreductases / antagonists & inhibitors
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NADH, NADPH Oxidoreductases / metabolism*
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Oxygen Consumption
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Phosphorylation / drug effects
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Protein Kinase Inhibitors
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Protein Kinases / pharmacology*
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Protein Serine-Threonine Kinases
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Pyrazines / metabolism
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Pyruvate Dehydrogenase Acetyl-Transferring Kinase
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Rats
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Rats, Wistar
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Reactive Oxygen Species / metabolism*
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Superoxides / metabolism
Substances
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Imidazoles
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Membrane Proteins
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Ndufa1 protein, mammalian
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Protein Kinase Inhibitors
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Pyrazines
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Pyruvate Dehydrogenase Acetyl-Transferring Kinase
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Reactive Oxygen Species
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NAD
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Superoxides
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coelenterazine
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Firefly Luciferin
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Adenosine Triphosphate
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Cyclic AMP
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NADH, NADPH Oxidoreductases
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NADH Dehydrogenase
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Protein Kinases
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Protein Serine-Threonine Kinases
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Cyclic AMP-Dependent Protein Kinases
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Electron Transport Complex I
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NDUFA1 protein, human
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NDUFS4 protein, human