N- and C-termini modulate the effects of pH and phosphorylation on hepatic 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase

Irwin J. Kurland, Brett Chapman, M. Raafat El-Maghrabi

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Liver and skeletal muscle isoforms of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (6PF2K/Fru-2,6-P2ase) isoenzymes are products of alternatively spliced first exons of the same gene, with common kinase and bisphosphatase domains. The muscle-specific exon-1 encodes nine unique amino acids, that lack the cAMP-dependent protein kinase (PK-A) phosphorylation site, and differ in sequence from those encoded by the liver-specific exon-1 (32 amino acids), contributing to its much lower affinity for fructose 6-phosphate (Fru-6-P). PK-A phosphorylation of the liver isoform at Ser32 reduces the affinity of the kinase for Fru-6-P, and stimulates the bisphosphatase V(max). In the present study, we have defined the locus of interaction of the N-terminal residues with the N-terminal kinase and C-terminal domains by successive N- and C-terminal deletions. This study shows that: (1) residues Gly5-Glu6-Leu7 of the liver isoform are responsible for increasing the affinity of 6PF2K for Fru-6-P, maintaining the inhibition of Fru-2,6-P2ase activity, and mediating the effects of PK-A phosphorylation on the two activities; (2) the loss of Fru-6-P inhibition of the bisphosphatase and the enhancement of its V(max), rather than the inhibition of the kinase, may be responsible for the behaviour of the muscle isoform primarily as a bisphosphatase; (3) the composition of residues 24-32 of the liver form appears to confer the enhanced kinase catalytic rate of this form over that of the muscle isoform. It is concluded that specific regions of the N-terminus of liver and skeletal muscle 6PF2K/Fru-2,6-P2ase have a role in adapting the two activities to work in the physiological range of pH and substrate concentrations found in each particular tissue.

Original languageEnglish (US)
Pages (from-to)459-467
Number of pages9
JournalBiochemical Journal
Volume347
Issue number2
DOIs
StatePublished - Apr 15 2000
Externally publishedYes

Fingerprint

Phosphofructokinase-2
Phosphorylation
Liver
Muscle
Protein Isoforms
Phosphotransferases
Exons
Muscles
Protein Kinases
Skeletal Muscle
Amino Acids
Cyclic AMP-Dependent Protein Kinases
Isoenzymes
Genes
Tissue
fructose-6-phosphate
Substrates
Chemical analysis

Keywords

  • Enzyme kinetics
  • Gluconeogenesis
  • Glycolysis
  • Hepatic glucose metabolism
  • Structure/function

ASJC Scopus subject areas

  • Biochemistry

Cite this

N- and C-termini modulate the effects of pH and phosphorylation on hepatic 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase. / Kurland, Irwin J.; Chapman, Brett; El-Maghrabi, M. Raafat.

In: Biochemical Journal, Vol. 347, No. 2, 15.04.2000, p. 459-467.

Research output: Contribution to journalArticle

@article{a7d03a0ebb644bb5bf73fc74ca56cb1b,
title = "N- and C-termini modulate the effects of pH and phosphorylation on hepatic 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase",
abstract = "Liver and skeletal muscle isoforms of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (6PF2K/Fru-2,6-P2ase) isoenzymes are products of alternatively spliced first exons of the same gene, with common kinase and bisphosphatase domains. The muscle-specific exon-1 encodes nine unique amino acids, that lack the cAMP-dependent protein kinase (PK-A) phosphorylation site, and differ in sequence from those encoded by the liver-specific exon-1 (32 amino acids), contributing to its much lower affinity for fructose 6-phosphate (Fru-6-P). PK-A phosphorylation of the liver isoform at Ser32 reduces the affinity of the kinase for Fru-6-P, and stimulates the bisphosphatase V(max). In the present study, we have defined the locus of interaction of the N-terminal residues with the N-terminal kinase and C-terminal domains by successive N- and C-terminal deletions. This study shows that: (1) residues Gly5-Glu6-Leu7 of the liver isoform are responsible for increasing the affinity of 6PF2K for Fru-6-P, maintaining the inhibition of Fru-2,6-P2ase activity, and mediating the effects of PK-A phosphorylation on the two activities; (2) the loss of Fru-6-P inhibition of the bisphosphatase and the enhancement of its V(max), rather than the inhibition of the kinase, may be responsible for the behaviour of the muscle isoform primarily as a bisphosphatase; (3) the composition of residues 24-32 of the liver form appears to confer the enhanced kinase catalytic rate of this form over that of the muscle isoform. It is concluded that specific regions of the N-terminus of liver and skeletal muscle 6PF2K/Fru-2,6-P2ase have a role in adapting the two activities to work in the physiological range of pH and substrate concentrations found in each particular tissue.",
keywords = "Enzyme kinetics, Gluconeogenesis, Glycolysis, Hepatic glucose metabolism, Structure/function",
author = "Kurland, {Irwin J.} and Brett Chapman and El-Maghrabi, {M. Raafat}",
year = "2000",
month = "4",
day = "15",
doi = "10.1042/0264-6021:3470459",
language = "English (US)",
volume = "347",
pages = "459--467",
journal = "Biochemical Journal",
issn = "0264-6021",
publisher = "Portland Press Ltd.",
number = "2",

}

TY - JOUR

T1 - N- and C-termini modulate the effects of pH and phosphorylation on hepatic 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase

AU - Kurland, Irwin J.

AU - Chapman, Brett

AU - El-Maghrabi, M. Raafat

PY - 2000/4/15

Y1 - 2000/4/15

N2 - Liver and skeletal muscle isoforms of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (6PF2K/Fru-2,6-P2ase) isoenzymes are products of alternatively spliced first exons of the same gene, with common kinase and bisphosphatase domains. The muscle-specific exon-1 encodes nine unique amino acids, that lack the cAMP-dependent protein kinase (PK-A) phosphorylation site, and differ in sequence from those encoded by the liver-specific exon-1 (32 amino acids), contributing to its much lower affinity for fructose 6-phosphate (Fru-6-P). PK-A phosphorylation of the liver isoform at Ser32 reduces the affinity of the kinase for Fru-6-P, and stimulates the bisphosphatase V(max). In the present study, we have defined the locus of interaction of the N-terminal residues with the N-terminal kinase and C-terminal domains by successive N- and C-terminal deletions. This study shows that: (1) residues Gly5-Glu6-Leu7 of the liver isoform are responsible for increasing the affinity of 6PF2K for Fru-6-P, maintaining the inhibition of Fru-2,6-P2ase activity, and mediating the effects of PK-A phosphorylation on the two activities; (2) the loss of Fru-6-P inhibition of the bisphosphatase and the enhancement of its V(max), rather than the inhibition of the kinase, may be responsible for the behaviour of the muscle isoform primarily as a bisphosphatase; (3) the composition of residues 24-32 of the liver form appears to confer the enhanced kinase catalytic rate of this form over that of the muscle isoform. It is concluded that specific regions of the N-terminus of liver and skeletal muscle 6PF2K/Fru-2,6-P2ase have a role in adapting the two activities to work in the physiological range of pH and substrate concentrations found in each particular tissue.

AB - Liver and skeletal muscle isoforms of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (6PF2K/Fru-2,6-P2ase) isoenzymes are products of alternatively spliced first exons of the same gene, with common kinase and bisphosphatase domains. The muscle-specific exon-1 encodes nine unique amino acids, that lack the cAMP-dependent protein kinase (PK-A) phosphorylation site, and differ in sequence from those encoded by the liver-specific exon-1 (32 amino acids), contributing to its much lower affinity for fructose 6-phosphate (Fru-6-P). PK-A phosphorylation of the liver isoform at Ser32 reduces the affinity of the kinase for Fru-6-P, and stimulates the bisphosphatase V(max). In the present study, we have defined the locus of interaction of the N-terminal residues with the N-terminal kinase and C-terminal domains by successive N- and C-terminal deletions. This study shows that: (1) residues Gly5-Glu6-Leu7 of the liver isoform are responsible for increasing the affinity of 6PF2K for Fru-6-P, maintaining the inhibition of Fru-2,6-P2ase activity, and mediating the effects of PK-A phosphorylation on the two activities; (2) the loss of Fru-6-P inhibition of the bisphosphatase and the enhancement of its V(max), rather than the inhibition of the kinase, may be responsible for the behaviour of the muscle isoform primarily as a bisphosphatase; (3) the composition of residues 24-32 of the liver form appears to confer the enhanced kinase catalytic rate of this form over that of the muscle isoform. It is concluded that specific regions of the N-terminus of liver and skeletal muscle 6PF2K/Fru-2,6-P2ase have a role in adapting the two activities to work in the physiological range of pH and substrate concentrations found in each particular tissue.

KW - Enzyme kinetics

KW - Gluconeogenesis

KW - Glycolysis

KW - Hepatic glucose metabolism

KW - Structure/function

UR - http://www.scopus.com/inward/record.url?scp=0034655718&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034655718&partnerID=8YFLogxK

U2 - 10.1042/0264-6021:3470459

DO - 10.1042/0264-6021:3470459

M3 - Article

C2 - 10749675

AN - SCOPUS:0034655718

VL - 347

SP - 459

EP - 467

JO - Biochemical Journal

JF - Biochemical Journal

SN - 0264-6021

IS - 2

ER -