Fructose 2,6-bisphosphate and AMP increase the affinity of the Ascaris suum phosphofructokinase for fructose 6-phosphate in a process separate from the relief of ATP inhibition.
Kinetic data have been collected suggesting that heterotropic activation by fructose 2,6-bisphosphate and AMP is a result not only of the relief of allosteric inhibition by ATP but is also the result of an increase in the affinity of phosphofructokinase for fructose 6-phosphate. Modification of the Ascaris suum phosphofructokinase at the ATP inhibitory site produces a form of the enzyme that no longer has hysteretic time courses or homotropic positive (fructose 6-phosphate) cooperativity or substrate inhibition (ATP) (Rao, G.S. J., Wariso, B.A., Cook, P.F., Hofer, H.W., and Harris, B.G. (1987a) J. Biol. Chem. 262, 14068-14073). This form of phosphofructokinase is Michaelis-Menten in its kinetic behavior but is still activated by fructose 2,6-bisphosphate and AMP and by phosphorylation using the catalytic subunit of cyclic AMP-dependent protein kinase (cAPK). Fructose 2,6-bisphosphate activates by decreasing KF-6-P by about 15-fold and has an activation constant of 92 nM, while AMP decreases KF-6-P about 6-fold and has an activation constant of 93 microM. Double activation experiments suggest that fructose 2,6-bisphosphate and AMP are synergistic in their activation. The desensitized form of the enzyme is phosphorylated by cAPK and has an increased affinity for fructose 6-phosphate in the absence of MgATP. The increased affinity results in a change in the order of addition of reactants from that with MgATP adding first for the nonphosphorylated enzyme to addition of fructose 6-phosphate first for the phosphorylated enzyme. The phosphorylated form of the enzyme is also still activated by fructose 2,6-bisphosphate and AMP.
Payne
M A
MA
Department of Microbiology and Immunology, Texas College of Osteopathic Medicine, Fort Worth 76107.
Rao
G S
GS
Harris
B G
BG
Cook
P F
PF
D000249:Adenosine Monophosphate / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000494:Allosteric Regulation; D000818:Animals; D001200:Ascaris / Q000201:enzymology*; D002942:Circular Dichroism; D004789:Enzyme Activation; D005635:Fructosediphosphates / Q000494:pharmacology*; D005636:Fructosephosphates / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D010732:Phosphofructokinase-1 / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000648:ultrastructure
D005635:Fructosediphosphates; D005636:Fructosephosphates; D000249:Adenosine Monophosphate; C027652:fructose 2,6-diphosphate; D000255:Adenosine Triphosphate; D010732:Phosphofructokinase-1