In Vitro Methods to Study AMPK

Abstract

AMPK studies in cell-free and cellular systems have significantly contributed to recent progress in the AMPK field. Biochemical characterization, structure determination and elucidation of AMPK-dependent signalling events benefit from application of state-of-the-art tools and methodology. This chapter provides a synopsis of recombinant protein expression systems and biochemical and cell-based study methods. We summarize three different expression systems for AMPK production: bacteria, insect cells and mammalian cells. In addition, kinase activity measurement, kinase substrate identification and determination of physical interaction with AMPK are discussed. The last part of this chapter focuses on the use of pharmacological activation, inhibition, and molecular genetic tools to study AMPK involvement in cellular signalling pathways.

Keywords: AMPK activity modulation; Genetic tools; Kinase activity determination; Recombinant protein expression; Substrate identification.