Ann Koay

Publications (5)19.74 Total impact

• Article: AMP-activated protein kinase β-subunit requires internal motion for optimal carbohydrate binding.

  Michael Bieri, Jesse I Mobbs, Ann Koay, Gavin Louey, Yee-Foong Mok, Danny M Hatters, Jong-Tae Park, Kwan-Hwa Park, Dietbert Neumann, David Stapleton, Paul R Gooley
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  ABSTRACT: AMP-activated protein kinase interacts with oligosaccharides and glycogen through the carbohydrate-binding module (CBM) containing the β-subunit, for which there are two isoforms (β(1) and β(2)). Muscle-specific β(2)-CBM, either as an isolated domain or in the intact enzyme, binds carbohydrates more tightly than the ubiquitous β(1)-CBM. Although residues that contact carbohydrate are strictly conserved, an additional threonine in a loop of β(2)-CBM is concurrent with an increase in flexibility in β(2)-CBM, which may account for the affinity differences between the two isoforms. In contrast to β(1)-CBM, unbound β(2)-CBM showed microsecond-to-millisecond motion at the base of a β-hairpin that contains residues that make critical contacts with carbohydrate. Upon binding to carbohydrate, similar microsecond-to-millisecond motion was observed in this β-hairpin and the loop that contains the threonine insertion. Deletion of the threonine from β(2)-CBM resulted in reduced carbohydrate affinity. Although motion was retained in the unbound state, a significant loss of motion was observed in the bound state of the β(2)-CBM mutant. Insertion of a threonine into the background of β(1)-CBM resulted in increased ligand affinity and flexibility in these loops when bound to carbohydrate. However, these mutations indicate that the additional threonine is not solely responsible for the differences in carbohydrate affinity and protein dynamics. Nevertheless, these results suggest that altered protein dynamics may contribute to differences in the ligand affinity of the two naturally occurring CBM isoforms.
  Biophysical Journal 01/2012; 102(2):305-14. · 3.65 Impact Factor
• Article: AMPK beta subunits display isoform specific affinities for carbohydrates.

  Ann Koay, Ben Woodcroft, Emma J Petrie, Helen Yue, Shane Emanuelle, Michael Bieri, Michael F Bailey, Mark Hargreaves, Jong-Tae Park, Kwan-Hwa Park, Stuart Ralph, Dietbert Neumann, David Stapleton, Paul R Gooley
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  ABSTRACT: AMP-activated protein kinase (AMPK) is a heterotrimer of catalytic (alpha) and regulatory (beta and gamma) subunits with at least two isoforms for each subunit. AMPK beta1 is widely expressed whilst AMPK beta2 is highly expressed in muscle and both beta isoforms contain a mid-molecule carbohydrate-binding module (beta-CBM). Here we show that beta2-CBM has evolved to contain a Thr insertion and increased affinity for glycogen mimetics with a preference for oligosaccharides containing a single alpha-1,6 branched residue. Deletion of Thr-101 reduces affinity for single alpha-1,6 branched oligosaccharides by 3-fold, while insertion of this residue into the equivalent position in the beta1-CBM sequence increases affinity by 3-fold, confirming the functional importance of this residue.
  FEBS letters 08/2010; 584(15):3499-503. · 3.54 Impact Factor
• Article: Thienopyridone drugs are selective activators of AMP-activated protein kinase beta1-containing complexes.

  John W Scott, Bryce J W van Denderen, Sebastian B Jorgensen, Jane E Honeyman, Gregory R Steinberg, Jonathan S Oakhill, Tristan J Iseli, Ann Koay, Paul R Gooley, David Stapleton, Bruce E Kemp
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  ABSTRACT: The AMP-activated protein kinase (AMPK) is an alphabetagamma heterotrimer that plays a pivotal role in regulating cellular and whole-body metabolism. Activation of AMPK reverses many of the metabolic defects associated with obesity and type 2 diabetes, and therefore AMPK is considered a promising target for drugs to treat these diseases. Recently, the thienopyridone A769662 has been reported to directly activate AMPK by an unexpected mechanism. Here we show that A769662 activates AMPK by a mechanism involving the beta subunit carbohydrate-binding module and residues from the gamma subunit but not the AMP-binding sites. Furthermore, A769662 exclusively activates AMPK heterotrimers containing the beta1 subunit. Our findings highlight the regulatory role played by the beta subunit in modulating AMPK activity and the possibility of developing isoform specific therapeutic activators of this important metabolic regulator.
  Chemistry & biology 12/2008; 15(11):1220-30. · 6.52 Impact Factor
• Article: AMP-activated protein kinase does not associate with glycogen alpha-particles from rat liver.

  Glendon J Parker, Ann Koay, Ryan Gilbert-Wilson, Lynne J Waddington, David Stapleton
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  ABSTRACT: The AMP-activated protein kinase (AMPK) is heterotrimer consisting of alpha catalytic subunit and beta/gamma regulatory subunits. It acts as a critical focal point for whole body and cellular mechanisms maintaining energy homeostasis by regulating carbohydrate and lipid metabolism, food intake, gene transcription, and protein synthesis. The AMPK beta subunit contains a glycogen-binding domain that has been shown to associate with glycogen particles in vitro and glycogen phosphorylase and glycogen synthase in cultured cells. To determine whether AMPK associates with glycogen particles in vivo, we developed a procedure to purify glycogen alpha-particles to apparent homogeneity from rat liver. Using immunoreactivity and mass spectrometry we determined that AMPK does not associate with the glycogen particle in livers from random-fed rats. This surprising finding indicates that the glycogen-binding properties of the AMPK beta subunit are likely regulated and responsive to the metabolic status of the hepatocyte.
  Biochemical and Biophysical Research Communications 12/2007; 362(4):811-5. · 2.48 Impact Factor
• Article: Oligosaccharide recognition and binding to the carbohydrate binding module of AMP-activated protein kinase.

  Ann Koay, Kieran A Rimmer, Haydyn D T Mertens, Paul R Gooley, David Stapleton
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  ABSTRACT: The AMP-activated protein kinase (AMPK) contains a carbohydrate-binding module (beta1-CBM) that is conserved from yeast to mammals. Beta1-CBM has been shown to localize AMPK to glycogen in intact cells and in vitro. Here we use Nuclear Magnetic Resonance spectroscopy to investigate oligosaccharide binding to 15N labelled beta1-CBM. We find that beta1-CBM shows greatest affinity to carbohydrates of greater than five glucose units joined via alpha,1-->4 glycosidic linkages with a single, but not multiple, glucose units in an alpha,1-->6 branch. The near identical chemical shift profile for all oligosaccharides whether cyclic or linear suggest a similar binding conformation and confirms the presence of a single carbohydrate-binding site.
  FEBS Letters 10/2007; 581(26):5055-9. · 3.54 Impact Factor