Title: Visualizing AMPK Drug Binding Sites Through Crystallization of Full-Length Phosphorylated alpha 2 beta 1 gamma 1 Heterotrimer
Authors: Langendorf, CG
Oakhill, JS
Kemp, BE
Issue Year: 2018
Publisher HUMANA PRESS INC
Series Methods Mol. Biol.:
Abstract Here, we describe the crystallization protocol for AMPK, including protein production and purification. AMPK can be readily crystallized in the presence of PEG to give diffracting crystals to a resolution of between 2.5 and 3.5 angstrom using synchrotron radiation. This method allows for visualization of drugs or small molecules that bind to the ADaM site, CBS sites, ATP binding site, and the newly identified C2 binding sites in the gamma-subunit via co-crystallization with phosphorylated AMPK (pT172) alpha 2 beta 1 gamma 1 isoform or alpha 2/1 beta 1 gamma 1 chimera. Drugs with binding affinities above 500 nM fail to co-crystallize with AMPK using these parameters.
URI: https://publications.svi.edu.au/publications/4709
ISSN
Other Identifiers 10.1007/978-1-4939-7598-3_2
Publication type Article