Covalent control of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase: Insights into autoregulation of a bifunctional enzyme

Irwin J. Kurland, S. J. Pilkis

Research output: Contribution to journalArticle

42 Citations (Scopus)

Abstract

The hepatic bifunctional enzyme, 6-phosphofructo-2-kinase/fructose-2,6 bisphosphatase (6PF-2-K/Fru-2,6-P2ase), E.C. 2-7-1-105/E.C. 3-1-3-46, is one member of a family of unique bifunctional proteins that catalyze the synthesis and degradation of the regulatory metabolite fructose-2,6- bisphosphate (Fru-2,6 P2). Fru-2,6-P2 is a potent activator of the glycolytic enzyme 6-phosphofructo-1-kinase and an inhibitor of the gluconeogenic enzyme fructose-1,6-bisphosphatase, and provides a switching mechanism between these two opposing pathways of hepatic carbohydrate metabolism. The activities of the hepatic 6PF-2-K/Fru-2,6-P2ase isoform are reciprocally regulated by a cyclic AMP-dependent protein kinase (cAPK) catalyzed phosphorylation at a single NH2-terminal residue, Ser-32. Phosphorylation at Ser-32 inhibits the kinase and activates the bisphosphatase, in part through an electrostatic mechanism. Substitution of Asp for Ser-32 mimics the effects of cAPK-catalyzed phosphorylation. In the dephosphorylated homodimer, the NH2- and COOH-terminal tail regions also have an interaction with their respective active sites on the same subunit to produce an autoregulatory inhibition of the bisphosphatase and activation of the kinase. In support of this hypothesis, deletion of either the NH2- or COOH-terminal tail region, or both regions, leads to a disruption of these interactions with a maximal activation of the bisphosphatase. Inhibition of the kinase is observed with the NH2-truncated forms, in which there is also a diminution of cAPK phosphorylation to decrease the K(m) for Fru-6-P. Phosphorylation of the bifunctional enzyme by cAPK disrupts these autoregulatory interactions, resulting in inhibition of the kinase and activation of the bisphosphatase. Therefore, effects of cyclic AMP-dependent phosphorylation are mediated by a combination of electrostatic and autoregulatory control mechanisms.

Original languageEnglish (US)
Pages (from-to)1023-1037
Number of pages15
JournalProtein Science
Volume4
Issue number6
StatePublished - 1995
Externally publishedYes

Fingerprint

Phosphofructokinase-2
Phosphorylation
Homeostasis
Cyclic AMP-Dependent Protein Kinases
Phosphotransferases
Enzymes
Chemical activation
Static Electricity
Tail
Electrostatics
Liver
Phosphofructokinase-1
Enzyme Activators
Fructose-Bisphosphatase
Carbohydrate Metabolism
Enzyme Inhibitors
Viperidae
Metabolites
Cyclic AMP
Catalytic Domain

Keywords

  • 6 phosphofructo-2-kinase/fructose-2,6-bisphosphatase
  • 6-phosphofructo-1- kinase inhibitor
  • bifunctional enzymes
  • cyclic AMP-dependent phosphorylation
  • fructose-1,6- bisphosphatase inhibitor
  • hepatic carbohydrate metabolism

ASJC Scopus subject areas

  • Biochemistry

Cite this

Covalent control of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase : Insights into autoregulation of a bifunctional enzyme. / Kurland, Irwin J.; Pilkis, S. J.

In: Protein Science, Vol. 4, No. 6, 1995, p. 1023-1037.

Research output: Contribution to journalArticle

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abstract = "The hepatic bifunctional enzyme, 6-phosphofructo-2-kinase/fructose-2,6 bisphosphatase (6PF-2-K/Fru-2,6-P2ase), E.C. 2-7-1-105/E.C. 3-1-3-46, is one member of a family of unique bifunctional proteins that catalyze the synthesis and degradation of the regulatory metabolite fructose-2,6- bisphosphate (Fru-2,6 P2). Fru-2,6-P2 is a potent activator of the glycolytic enzyme 6-phosphofructo-1-kinase and an inhibitor of the gluconeogenic enzyme fructose-1,6-bisphosphatase, and provides a switching mechanism between these two opposing pathways of hepatic carbohydrate metabolism. The activities of the hepatic 6PF-2-K/Fru-2,6-P2ase isoform are reciprocally regulated by a cyclic AMP-dependent protein kinase (cAPK) catalyzed phosphorylation at a single NH2-terminal residue, Ser-32. Phosphorylation at Ser-32 inhibits the kinase and activates the bisphosphatase, in part through an electrostatic mechanism. Substitution of Asp for Ser-32 mimics the effects of cAPK-catalyzed phosphorylation. In the dephosphorylated homodimer, the NH2- and COOH-terminal tail regions also have an interaction with their respective active sites on the same subunit to produce an autoregulatory inhibition of the bisphosphatase and activation of the kinase. In support of this hypothesis, deletion of either the NH2- or COOH-terminal tail region, or both regions, leads to a disruption of these interactions with a maximal activation of the bisphosphatase. Inhibition of the kinase is observed with the NH2-truncated forms, in which there is also a diminution of cAPK phosphorylation to decrease the K(m) for Fru-6-P. Phosphorylation of the bifunctional enzyme by cAPK disrupts these autoregulatory interactions, resulting in inhibition of the kinase and activation of the bisphosphatase. Therefore, effects of cyclic AMP-dependent phosphorylation are mediated by a combination of electrostatic and autoregulatory control mechanisms.",
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