Characterisation of 5'-AMP-activated protein kinase in human liver using specific peptide substrates and the effects of 5'-AMP analogues on enzyme activity.

A specific peptide (SAMS peptide) phosphorylation assay has previously been used to measure and subsequently purify rat liver 5'-AMP-activated protein kinase (AMPK). In this report, we show that this peptide and a peptide based on the sequence surrounding the site phosphorylated on 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase by AMPK (HMG peptide) can be used to measure human liver AMPK. Our data demonstrate that both human and rat AMPKs have a higher affinity for the HMG peptide compared to the SAMS peptide. We have used these peptide phosphorylation assays to identify novel activators of AMPK.