Homogenous Fluorescent Assays for Characterizing Small-Molecule Activators of AMP-Activated Protein Kinase (AMPK)
Laurie J Reichling, Steven M Riddle, Baigen Mei, Rica Bruinsma, Tony A Goossens, Kristin G Huwiler, Mark Maffitt, Alyssa M.G Newport, Xiao-Dong Qian‡, Carmen Ruttimann-Johnson, Kurt W Vogel*
Identifiers and Pagination:Year: 2008
First Page: 34
Last Page: 42
Publisher Id: CCGTM-1-34
Article History:Received Date: 29/12/2007
Revision Received Date: 15/1/2008
Acceptance Date: 21/1/2008
Electronic publication date: 25/2/2008
Collection year: 2008
open-access license: This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.5/), which permits unrestrictive use, distribution, and reproduction in any medium, provided the original work is properly cited.
AMP activated protein kinase (AMPK) is a key regulator of cellular metabolism. AMPK activity is modulated in part by binding of AMP to the γ-subunit of the kinase, which increases the activity of the catalytic α-subunit. Because increased AMPK activity in the liver and in skeletal muscle leads to increased fatty acid oxidation and decreased cholesterol and fatty acid biosynthesis, activators of AMPK are being sought for treatment of type-2 diabetes and other metabolic disorders. The unique mechanism of AMPK activation offers an opportunity to develop small molecules that directly upregulate AMPK activity, and there exists a need for simplified methods to identify and characterize small-molecules that show isoform-specific effects on AMPK. We have developed a suite of fluorescence-based assays to identify and characterize such compounds, and have used these to characterize and compare activity of recombinant AMPK α1β1γ1 and α2β1γ1 isoforms in response to small molecule activators and inhibitors.