Structural basis for AMP binding to mammalian AMP-activated protein kinase

Abstract

AMP-activated protein kinase (AMPK) regulates cellular metabolism in response to the availability of energy and is therefore a target for type II diabetes treatment. It senses changes in the ratio of AMP/ATP by binding both species in a competitive manner. Thus, increases in the concentration of AMP activate AMPK resulting in the phosphorylation and differential regulation of a series of downstream targets that control anabolic and catabolic pathways. We report here the crystal structure of the regulatory fragment of mammalian AMPK in complexes with AMP and ATP. The phosphate groups of AMP/ATP lie in a groove on the surface of the gamma domain, which is lined with basic residues, many of which are associated with disease-causing mutations. Structural and solution studies reveal that two sites on the gamma domain bind either AMP or Mg.ATP, whereas a third site contains a tightly bound AMP that does not exchange. Our binding studies indicate that under physiological conditions AMPK mainly exists in its inactive form in complex with Mg.ATP, which is much more abundant than AMP. Our modelling studies suggest how changes in the concentration of AMP ([AMP]) enhance AMPK activity levels. The structure also suggests a mechanism for propagating AMP/ATP signalling whereby a phosphorylated residue from the alpha and/or beta subunits binds to the gamma subunit in the presence of AMP but not when ATP is bound.

Ghrelin in obesity and endocrine diseases

“…AMPK is a regulator of cellular and systemic energy homeostasis and acts as a sensor of energy status (Kola et al, 2006;Xiao et al, 2007). Once activated, AMPK leads to a switch from anabolic to catabolic pathways; with the aim of restoring energy balance, ATP production is increased and ATP-utilising pathways are inhibited.…”

Adenosine Monophosphate-Activated Protein Kinase: A Central Regulator of Metabolism with Roles in Diabetes, Cancer, and Viral Infection

“…As in the original structure that lacked the kinase domain (Xiao et al 2007), the CTDs of a and b associate closely together, but in the new structure they also interact with the activation loop of the kinase domain, hindering access to Thr172 of protein phosphatases that would dephosphorylate the residue. The kinase domain and the a-CTD, at opposite ends of the a subunit, are connected by a long extended peptide that wraps around the g subunit, resembling two "arms" with the hands clasped together, holding the g subunit in a tight embrace (Fig.…”

“…ADP binding at site 3 also appears to promote phosphorylation of Thr172 by upstream kinases (Oakhill et al 2011). Site 4, whose function remains unclear, binds AMP very tightly; AMP is already present in this site when recombinant AMPK is purified from E. coli, and it does not exchange with ADP or ATP (Xiao et al 2007). …”

“…The structures of the core AMPK complexes from S. cerevisiae, S. pombe, and mammals show that the two Bateman domains formed by CBS1:CBS2 and CBS3:CBS4 assemble head to head to form a flattened disk with one CBS repeat in each quadrant (Amodeo et al 2007;Townley and Shapiro 2007;Xiao et al 2007). This symmetrical arrangement creates four clefts near the center where AMP, ADP, or ATP (referred to below as AXP) might bind, two accessible from one side of the disk and two from the other.…”

“…The carboxyterminal domain (CTD) of the b subunit wraps around this a-CTD and then terminates with two antiparallel strands. These form a b sheet with a third strand provided by the amino terminus of the g subunit; this b sheet forms the conserved "core" of the abg complex in S. cerevisiae, S. pombe, and mammals (Amodeo et al 2007;Townley and Shapiro 2007;Xiao et al 2007). Nearer to the center of the b subunit ( Fig.…”