6-phosphofructo 2-kinase/fructose 2,6-bisphosphatase in liver of the teleost Sparus aurata.
6-Phosphofructo 2-kinase/fructose 2,6-bisphosphatase was purified from the liver of the teleost fish Sparus aurata and the enzymatic activities were characterized kinetically. Both activities copurify, being dimers of relative molecular mass of 98 kDa with subunits of M(r) 54 kDa. Although both specific activities are in the range of mammalian liver isozymes, the Kmfru 6-P of teleost 6-phosphofructo 2-kinase is 3 times that in rat liver. The S. aurata 6-phosphofructo 2-kinase is inhibited by ADP, citrate and phosphoenolpyruvate, and fructose-2,6-bisphosphatase presents inhibition by fru 6-P. Unlike the rat liver enzyme, the kinase reaction is scarcely inhibited by glycerol 3-P. The teleost isozyme is substrate for the cyclic-AMP-dependent protein kinase, as can be followed by the incorporation of 32P from ATP into the enzyme. Phosphorylation of the enzyme changes its kinetic behavior, leading to a form with a lower kinase/bisphosphatase activity ratio. No change is detected in the fru 6-P dependence of 6-phosphofructo 2-kinase, but the phosphorylated form is more sensitive to inhibition by effectors, especially by glycerol 3-phosphate. Phosphorylation enhances the fructose-2,6-bisphosphatase Vmax activity twofold. The implications of all these kinetic characteristics in the control of hepatic fructose-2,6-bisphosphate levels are discussed in the context of the studies in S. aurata in vivo. The results support the hypothesis that differences in the regulation of 6-phosphofructo 2-kinase/fructose-2,6-bisphosphatase are a key point for the specific adaptations of carbohydrate metabolism in this teleost fish.
García de Frutos
P
P
Unitat de Bioquímica, Facultat de Farmàcia, Universitat de Barcelona, Spain.
Baanante
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IV
D000818:Animals; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D008322:Mammals; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D010473:Perciformes; D025481:Phosphofructokinase-2; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*
D007527:Isoenzymes; D046911:Macromolecular Substances; D009097:Multienzyme Complexes; D017853:Phosphotransferases (Alcohol Group Acceptor); D025481:Phosphofructokinase-2; D010744:Phosphoric Monoester Hydrolases