Kinase activity determination of specific AMPK complexes/heterotrimers in the skeletal muscle

Abstract

Measuring the kinase activity of the 5′-AMP-activated protein kinase (AMPK) is an essential part of understanding the regulation of this metabolic master switch. The AMPK heterotrimer can exist in 12 different constellations with potentially diverse activation patterns. It is therefore important to be able to measure heterotrimer-specific activity to discriminate between these patterns. In this chapter we describe how to measure the AMPK activity of specific heterotrimeric complexes by consecutive immunoprecipitations and how the assay can be performed in a medium throughput fashion using 96-well plates.