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The glycogen-binding domain on the AMPK beta subunit allows the kinase to act as a glycogen sensor.

McBride A, Ghilagaber S, Nikolaev A, Hardie DG - Cell Metab. (2009)

Bottom Line: We synthesized a series of branched oligosaccharides and show that those with a single alpha1-->6 branch are allosteric inhibitors that also inhibit phosphorylation by upstream kinases.Inhibition by all carbohydrates tested was dependent on the glycogen-binding domain being abolished by mutation of residues required for carbohydrate binding.Our results suggest the hypothesis that AMPK, as well as monitoring immediate energy availability by sensing AMP/ATP, may also be able to sense the status of cellular energy reserves in the form of glycogen.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Physiology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, UK.

ABSTRACT
AMPK beta subunits contain a conserved domain that causes association with glycogen. Although glycogen availability is known to affect AMPK regulation in vivo, the molecular mechanism for this has not been clear. We now show that AMPK is inhibited by glycogen, particularly preparations with high branching content. We synthesized a series of branched oligosaccharides and show that those with a single alpha1-->6 branch are allosteric inhibitors that also inhibit phosphorylation by upstream kinases. Removal of the outer chains of glycogen using phosphorylase, thus exposing the outer branches, renders inhibition of AMPK more potent. Inhibition by all carbohydrates tested was dependent on the glycogen-binding domain being abolished by mutation of residues required for carbohydrate binding. Our results suggest the hypothesis that AMPK, as well as monitoring immediate energy availability by sensing AMP/ATP, may also be able to sense the status of cellular energy reserves in the form of glycogen.

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Related in: MedlinePlus

Inhibition of Rat Liver AMPK by Synthetic Branched Oligosaccharides(A) Table showing the identification (ID) number, structure, and estimated IC50 ± SEM.(B) Inhibition of purified rat liver AMPK by oligosaccharides. These data were used to generate the IC50 values shown in (A), and the curves were drawn using the best-fit parameters. A key to the ID of each oligosaccharide is shown on the right.(C) Inhibition of recombinant wild-type and W100G/W133A mutant AMPK (α1β1γ1 complex) by synthetic oligosaccharides, numbered as in (A) and (B). The data are presented with the oligosaccharides in order of increasing potency from left to right; results are mean ± SEM (n = 3). Significant differences ± oligosaccharide for the wild-type by two-way ANOVA. ∗p < 0.05; ∗∗∗p < 0.001.
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fig6: Inhibition of Rat Liver AMPK by Synthetic Branched Oligosaccharides(A) Table showing the identification (ID) number, structure, and estimated IC50 ± SEM.(B) Inhibition of purified rat liver AMPK by oligosaccharides. These data were used to generate the IC50 values shown in (A), and the curves were drawn using the best-fit parameters. A key to the ID of each oligosaccharide is shown on the right.(C) Inhibition of recombinant wild-type and W100G/W133A mutant AMPK (α1β1γ1 complex) by synthetic oligosaccharides, numbered as in (A) and (B). The data are presented with the oligosaccharides in order of increasing potency from left to right; results are mean ± SEM (n = 3). Significant differences ± oligosaccharide for the wild-type by two-way ANOVA. ∗p < 0.05; ∗∗∗p < 0.001.

Mentions: To test whether oligosaccharides larger than isomaltose with an α1→6 linkage would be more potent inhibitors, we chemically synthesized a series of α1→4 linked oligosaccharides, containing from three to six glucose units, with a single α1→6 linkage (Figure 6A). Because oligosaccharides with a free reducing end exist in solution as mixtures of α and β anomers, these oligosaccharides were synthesized with a methoxy group at carbon 1 of the glucose at the reducing end, in the α configuration. For comparison, we also synthesized methyl α-maltoside and methyl α-isomaltoside.


The glycogen-binding domain on the AMPK beta subunit allows the kinase to act as a glycogen sensor.

McBride A, Ghilagaber S, Nikolaev A, Hardie DG - Cell Metab. (2009)

Inhibition of Rat Liver AMPK by Synthetic Branched Oligosaccharides(A) Table showing the identification (ID) number, structure, and estimated IC50 ± SEM.(B) Inhibition of purified rat liver AMPK by oligosaccharides. These data were used to generate the IC50 values shown in (A), and the curves were drawn using the best-fit parameters. A key to the ID of each oligosaccharide is shown on the right.(C) Inhibition of recombinant wild-type and W100G/W133A mutant AMPK (α1β1γ1 complex) by synthetic oligosaccharides, numbered as in (A) and (B). The data are presented with the oligosaccharides in order of increasing potency from left to right; results are mean ± SEM (n = 3). Significant differences ± oligosaccharide for the wild-type by two-way ANOVA. ∗p < 0.05; ∗∗∗p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2642990&req=5

fig6: Inhibition of Rat Liver AMPK by Synthetic Branched Oligosaccharides(A) Table showing the identification (ID) number, structure, and estimated IC50 ± SEM.(B) Inhibition of purified rat liver AMPK by oligosaccharides. These data were used to generate the IC50 values shown in (A), and the curves were drawn using the best-fit parameters. A key to the ID of each oligosaccharide is shown on the right.(C) Inhibition of recombinant wild-type and W100G/W133A mutant AMPK (α1β1γ1 complex) by synthetic oligosaccharides, numbered as in (A) and (B). The data are presented with the oligosaccharides in order of increasing potency from left to right; results are mean ± SEM (n = 3). Significant differences ± oligosaccharide for the wild-type by two-way ANOVA. ∗p < 0.05; ∗∗∗p < 0.001.
Mentions: To test whether oligosaccharides larger than isomaltose with an α1→6 linkage would be more potent inhibitors, we chemically synthesized a series of α1→4 linked oligosaccharides, containing from three to six glucose units, with a single α1→6 linkage (Figure 6A). Because oligosaccharides with a free reducing end exist in solution as mixtures of α and β anomers, these oligosaccharides were synthesized with a methoxy group at carbon 1 of the glucose at the reducing end, in the α configuration. For comparison, we also synthesized methyl α-maltoside and methyl α-isomaltoside.

Bottom Line: We synthesized a series of branched oligosaccharides and show that those with a single alpha1-->6 branch are allosteric inhibitors that also inhibit phosphorylation by upstream kinases.Inhibition by all carbohydrates tested was dependent on the glycogen-binding domain being abolished by mutation of residues required for carbohydrate binding.Our results suggest the hypothesis that AMPK, as well as monitoring immediate energy availability by sensing AMP/ATP, may also be able to sense the status of cellular energy reserves in the form of glycogen.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Physiology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, UK.

ABSTRACT
AMPK beta subunits contain a conserved domain that causes association with glycogen. Although glycogen availability is known to affect AMPK regulation in vivo, the molecular mechanism for this has not been clear. We now show that AMPK is inhibited by glycogen, particularly preparations with high branching content. We synthesized a series of branched oligosaccharides and show that those with a single alpha1-->6 branch are allosteric inhibitors that also inhibit phosphorylation by upstream kinases. Removal of the outer chains of glycogen using phosphorylase, thus exposing the outer branches, renders inhibition of AMPK more potent. Inhibition by all carbohydrates tested was dependent on the glycogen-binding domain being abolished by mutation of residues required for carbohydrate binding. Our results suggest the hypothesis that AMPK, as well as monitoring immediate energy availability by sensing AMP/ATP, may also be able to sense the status of cellular energy reserves in the form of glycogen.

Show MeSH
Related in: MedlinePlus