Kinase assays and predicted phosphorylation sequence of representative AMPK targets. (A) AMPK phosphorylation consensus sequence. Blue represents basic amino acids, orange represents hydrophobic amino acids, and red represents phosphorylated amino acids. (B-D) Coomassie blue staining (CB) and autoradiography of in vitro kinase assays without (left) and with activated AMPK (right), (γ-32P) ATP, and full-length recombinant substrates. Arrow (→) indicates substrate. Each panel is accompanied by the control kinase assay that contained only recombinant, active AMPK and (γ-32P) ATP to identify relative position of autophosphorylated AMPK subunits.