title: Regulation of 6-phosphofructo-2-kinase activity by cyclic AMP-dependent phosphorylation.
abstract: Addition of glucagon to isolated rat hepatocytes resulted in inhibition of 6-phosphofructo-2-kinase (ATP:D-fructose-6-phosphate-2-phosphotransferase) activity in extracts of the cells and in a decrease in the intracellular level of fructose 2,6-bisphosphate. The effect on 6-phosphofructo-2-kinase was characterized by a decrease in the affinity of the enzyme for fructose 6-phosphate. To investigate the mechanism of action of glucagon, 6-phosphofructo-2-kinase from rat liver was partially purified by polyethylene glycol precipitation, DEAE-cellulose chromatography, (NH4)2SO4 fractionation, Sephacryl S-200 gel filtration, DEAE-Sephadex chromatography, and Sephadex G-100 gel filtration. Incubation of the purified enzyme with the catalytic subunit of the cyclic AMP-dependent protein kinase from rat liver and [gamma-32P]ATP resulted in 32P incorporation into a protein with a subunit Mr of 49,000 as determined by NaDodSO4 disc gel electrophoresis. Associated with this phosphorylation was an inhibition of 6-phosphofructo-2-kinase activity that was also characterized by a decrease in the affinity of the enzyme for fructose-6-phosphate. Both the phosphorylation and the inhibition of the purified 6-phosphofructo-2-kinase were blocked by addition of the heat-stable protein kinase inhibitor. It is concluded that the glucagon-induced decrease in fructose 2,6-bisphosphate levels observed in isolated hepatocytes is due, at least in part, to cyclic AMP-dependent phosphorylation and inhibition of 6-phosphofructo-2-kinase.
mesh_terms: D000818:Animals; D000242:Cyclic AMP / Q000378:metabolism*; D004789:Enzyme Activation; D005635:Fructosediphosphates / Q000378:metabolism*; D005934:Glucagon / Q000494:pharmacology; D006598:Hexosediphosphates / Q000378:metabolism*; D008099:Liver / Q000201:enzymology; D008297:Male; D010732:Phosphofructokinase-1 / Q000302:isolation & purification / Q000378:metabolism; D025481:Phosphofructokinase-2; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D013329:Structure-Activity Relationship
chemical_list: D005635:Fructosediphosphates; D006598:Hexosediphosphates; D005934:Glucagon; D000242:Cyclic AMP; D010770:Phosphotransferases; D011494:Protein Kinases; D025481:Phosphofructokinase-2; D010732:Phosphofructokinase-1