Phosphate dependency of phosphofructokinase 2.
Experiments performed at micromolar concentrations of inorganic phosphate support the conclusion that liver phosphofructokinase 2 would be completely inactive in the absence of inorganic phosphate or arsenate. The concentration of inorganic phosphate that allowed half-maximal activity decreased with increasing pH, being approximately 0.11 mM at pH 6.5 and 0.05 mM at pH 8. The effect of phosphate was to increase V and to decrease Km for fructose 6-phosphate, without affecting Km for ATP. Citrate and P-enolpyruvate inhibited the enzyme non-competitively with fructose 6-phosphate and independently of the concentration of inorganic phosphate. Phosphorylation of the enzyme by the catalytic subunit of cyclic-AMP-dependent protein kinase did not markedly modify the phosphate requirement and its effect of inactivating phosphofructokinase 2 could not be counteracted by excess phosphate. A nearly complete phosphate dependency was also observed with phosphofructokinase 2 purified from Saccharomyces cerevisiae or from spinach leaves. By contrast, the fructose 2,6-bisphosphatase activity of the liver bifunctional enzyme was not dependent on the presence of inorganic phosphate. Phosphate increased this activity about threefold when measured in the absence of added fructose 6-phosphate and a half-maximal effect was reached at approximately 0.5 mM phosphate. Like glycerol phosphate, phosphate counteracted the inhibition of fructose 2,6-bisphosphatase by fructose 6-phosphate, but a much higher concentration of phosphate than of glycerol phosphate was required to reach this effect.
Laloux
M
M
Van Schaftingen
E
E
Francois
J
J
Hers
H G
HG
D000249:Adenosine Monophosphate / Q000494:pharmacology; D001149:Arsenates / Q000494:pharmacology; D002951:Citrates / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D005635:Fructosediphosphates / Q000494:pharmacology; D008099:Liver / Q000201:enzymology; D008832:Microchemistry; D010710:Phosphates / Q000494:pharmacology*; D010728:Phosphoenolpyruvate / Q000494:pharmacology; D025481:Phosphofructokinase-2; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors; D010770:Phosphotransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010944:Plants / Q000201:enzymology; D012441:Saccharomyces cerevisiae / Q000201:enzymology
D001149:Arsenates; D002951:Citrates; D005635:Fructosediphosphates; D010710:Phosphates; D000249:Adenosine Monophosphate; D010728:Phosphoenolpyruvate; C027652:fructose 2,6-diphosphate; D010770:Phosphotransferases; D025481:Phosphofructokinase-2; D010744:Phosphoric Monoester Hydrolases; C025657:arsenic acid