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Fig. 1. TAP-purification of AMPK-related kinases. (A) The pEGFP-C2-TAP vector, depicting the affinity tag added to the N-terminus of AMPK{alpha}1 and the AMPK-related kinases, consisting of a green fluorescent protein (GFP), protein A and a calmodulin-binding motif (CAM) flanked by TEV and precision protease cleavage sites, prior to multiple cloning sites. (B) Outline of the strategy that was employed to affinity purify the GFP-TAP-tagged kinases. (C) The TAP affinity purified kinases were electrophoresed on a polyacrylamide gel and the protein bands visualized following colloidal Coomassie Blue staining. Each of the major bands observed that we were able to identify by mass spectrometry was numbered and its identity indicated in Table 1. Bands that were not reliably identified are left unlabelled. *The bands identified as the bait kinase.





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