Structural properties of AMP-activated protein kinase : Dimerization, molecular shape, and changes upon ligand binding

Riek, Uwe and Scholz, Roland and Konarev, Peter and Rufer, Arne and Suter, Marianne and Nazabal, Alexis and Ringler, Philippe and Chami, Mohamed and Müller Shirley, A. and Neumann, Dietbert and Forstner, Michael and Hennig, Michael and Zenobi, Renato and Engel, Andreas and Svergun, Dmitri and Schlattner, Uwe and Wallimann, Theo. (2008) Structural properties of AMP-activated protein kinase : Dimerization, molecular shape, and changes upon ligand binding. Journal of biological chemistry, Vol. 283, H. 26. pp. 18331-18343.

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Official URL: http://edoc.unibas.ch/dok/A5262464

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Heterotrimeric AMP-activated protein kinase (AMPK) is crucial for energy homeostasis of eukaryotic cells and organisms. Here we report on (i) bacterial expression of untagged mammalian AMPK isoform combinations, all containing gamma(1), (ii) an automated four-dimensional purification protocol, and (iii) biophysical characterization of AMPK heterotrimers by small angle x-ray scattering in solution (SAXS), transmission and scanning transmission electron microscopy (TEM, STEM), and mass spectrometry (MS). AMPK in solution at low concentrations (~1 mg/ml) largely consisted of individual heterotrimers in TEM analysis, revealed a precise 1:1:1 stoichiometry of the three subunits in MS, and behaved as an ideal solution in SAXS. At higher AMPK concentrations, SAXS revealed concentration-dependent, reversible dimerization of AMPK heterotrimers and formation of higher oligomers, also confirmed by STEM mass measurements. Single particle reconstruction and averaging by SAXS and TEM, respectively, revealed similar elongated, flat AMPK particles with protrusions and an indentation. In the lower AMPK concentration range, addition of AMP resulted in a significant decrease of the radius of gyration by approximately 5% in SAXS, which indicates a conformational switch in AMPK induced by ligand binding. We propose a structural model involving a ligand-induced relative movement of the kinase domain resulting in a more compact heterotrimer and a conformational change in the kinase domain that protects AMPK from dephosphorylation of Thr(172), thus positively affecting AMPK activity.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Former Organization Units Biozentrum > Structural Biology (Engel)
UniBasel Contributors:Müller, Shirley and Chami, Mohamed and Engel, Andreas H and Ringler, Philippe
Item Type:Article, refereed
Article Subtype:Research Article
Publisher:American Society of Biological Chemists
Note:Publication type according to Uni Basel Research Database: Journal article
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Last Modified:22 Mar 2012 14:21
Deposited On:22 Mar 2012 13:24

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