Science for Health
09 January 2014
Researchers at NIMR have identified the drug binding site in an enzyme that regulates glucose, lipid and cholesterol metabolism. The research is published in Nature Communications.
The AMP-activated protein kinase (AMPK) enzyme is an evolutionarily conserved metabolic sensor found in all eukaryotes. Its primary function is to maintain energy homeostasis and cell survival. AMPK does this by integrating nutrient availability, environmental signals and energy expenditure. AMPK has attracted widespread attention as a potential drug target for the treatment of metabolic disorders such as the metabolic syndrome - a disorder of energy utilisation and storage.
AMPK is a heterotrimer consisting of a catalytic α, and regulatory β and γ subunits. AMPK is activated when the ATP levels within the cell decrease, and AMP/ADP concentrations increase. Steve Gamblin’s group, in NIMR’s Division of Molecular Structure, have previously shown that regulation by nucleotides is achieved through binding to two sites in the γ subunit. In 2006, Abbott Laboratories developed the first direct AMPK activator and showed that this compound, A-769662, displays beneficial effects in treating aspects of the metabolic syndrome. Consequently, there has been a major effort to determine the mechanism for activation of AMPK by A-769662 and to identify other AMPK activators that perform better than A-769662. Despite numerous studies, the identity of the drug binding site that regulates AMPK activity has remained elusive.
Using biochemical and biophysical approaches Steve Gamblin (pictured) and his group have now identified an activator of AMPK, termed 991, that was more potent than A-769662 at binding to and activating AMPK. A crystal structure of the full-length human AMPK complex was determined in the presence of 991 or A-769662. Their work showed that there is a single site for drug binding at an interface generated by the kinase domain (α subunit) and carbohydrate-binding module (CBM) (β subunit).
This work has revealed the drug binding site on AMPK for the first time and shows that 991/A-769662 activation of AMPK is achieved through binding to a different site than that used by AMPK’s physiological regulators, AMP, ADP and ATP.
This work is a major step forward in the AMPK field since it has implications for the design of potent activators of AMPK that can be used to treat metabolic disorders. Furthermore, it reveals the location of the CBM within the AMPK complex.
Inspection of the interface, with the activator removed reveals that the protein subunits make contacts around the rim of the interface but leave a large, significantly hydrophobic, cavity where the activator binds. ‘This raises the possibility that a natural metabolite may bind to this site and regulate AMPK. This could represent an important missing link in our understanding of the physiological regulation of AMPK.
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(a) Bar diagram indicating the three subunits that make up the complex.
(b) Cartoon representation of full-length α2β1γ1 in complex with the activator 991, the domains of the three subunits are coloured according to (a).
(c) (d) (e) Detailed view of 991 binding in a pocket generated at the interface between the CBM and the kinase domain
Bing Xiao, Matthew J. Sanders, David Carmena, Nicola J. Bright, Lesley F. Haire, Elizabeth Underwood, Bhakti R. Patel, Richard B. Heath, Philip A. Walker, Stefan Hallen, Fabrizio Giordanetto, Stephen R. Martin, David Carling & Steven J. Gamblin (2013)
Structural basis of AMPK regulation by small molecule activators
Nature Communications 4: 3017. Article fulltext
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