title: Reversible phosphorylation of pyruvate dehydrogenase in rat skeletal-muscle mitochondria. Effects of starvation and diabetes.
abstract: The total activity of pyruvate dehydrogenase (PDH) complex in rat hind-limb muscle mitochondria was 76.4 units/g of mitochondrial protein. The proportion of complex in the active form was 34% (as isolated), 8-14% (incubation with respiratory substrates) and greater than 98% (incubation without respiratory substrates). Complex was also inactivated by ATP in the presence of oligomycin B and carbonyl cyanide m-chlorophenylhydrazone. Ca2+ (which activates PDH phosphatase) and pyruvate or dichloroacetate (which inhibit PDH kinase) each increased the concentration of active PDH complex in a concentration-dependent manner in mitochondria oxidizing 2-oxoglutarate/L-malate. Values giving half-maximal activation were 10 nM-Ca2+, 3 mM-pyruvate and 16 microM-dichloroacetate. Activation by Ca2+ was inhibited by Na+ and Mg2+. Mitochondria incubated with [32P]Pi/2-oxoglutarate/L-malate incorporated 32P into three phosphorylation sites in the alpha-chain of PDH; relative rates of phosphorylation were sites 1 greater than 2 greater than 3, and of dephosphorylation, sites 2 greater than 1 greater than 3. Starvation ( 48h ) or induction of alloxan-diabetes had no effect on the total activity of PDH complex in skeletal-muscle mitochondria, but each decreased the concentration of active complex in mitochondria oxidizing 2-oxoglutarate/L-malate and increased the concentrations of Ca2+, pyruvate or dichloracetate required for half-maximal reactivation. In extracts of mitochondria the activity of PDH kinase was increased 2-3-fold by 48 h starvation or alloxan-diabetes, but the activity of PDH phosphatase was unchanged.
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology; D003999:Dichloroacetic Acid / Q000494:pharmacology; D004789:Enzyme Activation; D066298:In Vitro Techniques; D008931:Mitochondria, Muscle / Q000187:drug effects / Q000201:enzymology*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011769:Pyruvate Dehydrogenase (Lipoamide)-Phosphatase / Q000378:metabolism; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism*; D011773:Pyruvates / Q000494:pharmacology; D019289:Pyruvic Acid; D051381:Rats; D011919:Rats, Inbred Strains; D013217:Starvation / Q000201:enzymology
chemical_list: D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011768:Pyruvate Dehydrogenase Complex; D011773:Pyruvates; D019289:Pyruvic Acid; D000255:Adenosine Triphosphate; D003999:Dichloroacetic Acid; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011769:Pyruvate Dehydrogenase (Lipoamide)-Phosphatase; D002118:Calcium