Regulation of NADH/CoQ oxidoreductase: do phosphorylation events affect activity?
We had previously suggested that phosphorylation of proteins by mitochondrial kinases regulate the activity of NADH/CoQ oxidoreductase. Initial data showed that pyruvate dehydrogenase kinase (PDK) and cAMP-dependent protein kinase A (PKA) phosphorylate mitochondrial membrane proteins. Upon phosphorylation with crude PDK, mitochondria appeared to be deficient in NADH/cytochrome c reductase activity associated with increased superoxide production. Conversely, phosphorylation by PKA resulted in increased NADH/cytochrome c reductase activity and decreased superoxide formation. Current data confirms PKA involvement in regulating Complex I activity through phosphorylation of an 18 kDa subunit. Beef heart NADH/ cytochrome c reductase activity increases to 150% of control upon incubation with PKA and ATP-gamma-S. We have cloned the four human isoforms of PDK and purified beef heart Complex I. Incubation of mitochondria with PDK isoforms and ATP did not alter Complex I activity or superoxide production. Radiolabeling of mitochondria and purified Complex I with PDK failed to reveal phosphorylated proteins.
Maj
Mary C
MC
Metabolic Research Programme, The Hospital for Sick Children, 555 University Avenue, Toronto M5G 1X8, Canada.
Raha
Sandeep
S
Myint
Tomoko
T
Robinson
Brian H
BH
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D017868:Cyclic AMP-Dependent Protein Kinases; D042967:Electron Transport Complex I / Q000302:isolation & purification / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007658:Ketone Oxidoreductases / Q000378:metabolism*; D008929:Mitochondria, Heart / Q000201:enzymology*; D008969:Molecular Sequence Data; D009243:NAD / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D013481:Superoxides / Q000378:metabolism*; D013552:Swine
D007527:Isoenzymes; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011994:Recombinant Proteins; D009243:NAD; D013481:Superoxides; D000255:Adenosine Triphosphate; D007658:Ketone Oxidoreductases; C053164:pyruvate dehydrogenase (NADP+); D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D042967:Electron Transport Complex I