Significance of phosphorylation of phosphofructokinase (original) (raw)
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Biochemical and Biophysical Research Communications, 1988
Citrate, ATP and AMP affect similarly the activation of muscle phosphofructokinase by a-glucose 1,6-bisphosphate and fructose 1,6bisphosphate, but they affect differently its activation by fructose 2,6-bisphosphate. Activation by a-glucose 1,6-bisphosphate and fructose 2,6-bisphosphate is also differently affected by pH. This suggest that beth u-glucose 1,6-bisphosphate and fructose 1,6-bisphosphate induce thesame conformational change on muscle phosphofructokinase, distinct fromthat produced by fructose 2,6-bisphosphate.
Journal of Biological Chemistry
The effect of natural "activation factor" and synthetic fructose-2,6-Pz on the allosteric kinetic properties of liver and muscle phosphofructokinases was investigated. Both synthetic and natural fructose-2,6-Pz show identical effects on the allosteric kinetic properties of both enzymes. Auctose-2,6-Pz counteracts inhibition by ATP and citrate and decreases the K,,, for fructose-6-P. This fructose ester also acts synergistically with AMP in releasing ATP inhibition. The K,,, values of liver and muscle phosphofructokinase for fructose-2,6-Pz in the presence of 1.26 IMI ATP are 12 milliunits/ml (or 24 l ũ r ) and 5 milliunits/ml (or 10 m), respectively. At near physiological concentrations of ATP (3 1 1 1 ) and fructose-6-P (0.2 m), however, the K,,, values for fructose-2,6-Pz are increased to 12 PM and 0.8 p t for liver and muscle enzymes, respectively. Thus, fructose-2,6-Pz is the most potent activator of the enzyme compared to other known activators such as fructose-1,6-Pz. The rates of the reaction catalyzed by the enzymes under the above conditions are nonlinear: the rates decelerate in the absence or in the presence of lower concentrations of fructose-2,6-Pz, but the rates become linear in the presence of higher concentrations of fructose-2,6-Pz. Fructose-2,6-Pz also protects phosphofructokinase against inactivation by heat. Fructose-2,6-Pz, therefore, may be the most important allosteric effector in regulation of phosphofructokinase in liver as well as in other tissues.
Journal of Biological Chemistry
Low phosphate and high phosphate forms of phosphofructokinase (Furuya, E., and Uyeda, K. (1980) J. Biol. Chem. 255,[11656][11657][11658][11659] from rat liver were purified to homogeneity and various properties were compared. The specific activities of these enzymes and their electrophoretic mobilities on polyacrylamide in sodium dodecyl sulfate are the same. A limited tryptic digestion yields products with no change in the enzyme activity but with a reduction in the molecular weight of about 2000. Both low and high phosphate enzymes can be phosphorylated by the catalytic subunit of CAMP-dependent protein kinase, and approximately twice as much [32P]phosphate is incorporated into the low phosphate than the high phosphate enzyme.
Chicken liver phosphofructokinase. III. Kinetics and allosteric properties
Journal of Biological Chemistry
Some kinetic and allosteric properties of crystalline chicken liver phosphofructokinase were investigated. Lineweaver-Burk plots with fructose 6.phosphate and ATP as substrates yield a series of apparently parallel lines. The liver enzyme also catalyzes the phosphorylation of fructose-l-P, and the double reciprocal plot with fructose-l-P as the substrate yields intersecting lines. Phosphorylation of both fructose-6-P and fructose-l-P are lost at 4" at equal rate. Of the many sugar phosphates examined, only glucose-6-P, 6-P.
Journal of Biological Chemistry
Liver phosphofructokinase has been separated into three fractions by DEAE-cellulose chromatography. Chromatography of 32P-labeled enzyme reveals that the first fraction contains an average of 1.2 mol of phosphate/mol of enzyme (320,000 daltons), while the second and third fractions contain 3.3 mol of phosphate/ mol of enzyme. The high phosphate forms are much more sensitive to ATP inhibition than the low phosphate form.
Interaction of immobilized phosphofructokinase with soluble muscle proteins
Biochimica et Biophysica Acta (BBA) - General Subjects, 1986
Selected glycolytic enzymes (including phosphoglucose isomerase, aldolase, glyceraldehyde phosphate dehydrogenase, enolase, pyruvate kinase and lactate dehydrogenase), as well as glycogen phosphorylase, creatine kinase, and adenylate kinase, bound to phosphofructokinase immobilized on an agarose gel. The affinity of phosphofructokinase to these various proteins differed, with phosphorylase exhibiting the strongest binding. Binding was reversed either by: (1) elution with high-ionic-strength buffer (0.4 M KCI); (2) the addition of a 5-10 mM concentration of ATP; or (3) high concentrations of fructose 6-phosphate (5 mM).
An activation factor of liver phosphofructokinase
Proceedings of the National Academy of Sciences, 1980
Pure phosphofructokinase (ATP:D-fructose-6-phosphate 1-phosphotransferase, EC 2.7.1.11) from liver is strongly inhibited by ATP, whereas crude phosphofructokinase is only slightly inhibited by ATP. A factor that is removed from the enzyme during purification and can prevent the inhibition of phosphofructokinase by ATP has been isolated. The factor can be resolved into three components that differ in molecular weights, as shown by gel filtration on Sephadex G-25. These factors overcome the ATP inhibition but have no effect on the catalytic activity under the optimum assay conditions. Furthermore, AMP acts syngeristically with the activation factor in reversing ATP inhibition. It is proposed that the activation of phosphofructokinase by the activation factor and AMP is sufficient to account for the glycolytic flux in the liver.