title: A revised model for AMP-activated protein kinase structure: The alpha-subunit binds to both the beta- and gamma-subunits although there is no direct binding between the beta- and gamma-subunits.
abstract: The 5'-AMP-activated protein kinase (AMPK) is a master sensor for cellular metabolic energy state. It is activated by a high AMP/ATP ratio and leads to metabolic changes that conserve energy and utilize alternative cellular fuel sources. The kinase is composed of a heterotrimeric protein complex containing a catalytic alpha-subunit, an AMP-binding gamma-subunit, and a scaffolding beta-subunit thought to bind directly both the alpha- and gamma-subunits. Here, we use coimmunoprecipitation of proteins in transiently transfected cells to show that the alpha2-subunit binds directly not only to the beta-subunit, confirming previous work, but also to the gamma1-subunit. Deletion analysis of the alpha2-subunit reveals that the C-terminal 386-552 residues are sufficient to bind to the beta-subunit. The gamma1-subunit binds directly to the alpha2-subunit at two interaction sites, one within the catalytic domain consisting of alpha2 amino acids 1-312 and a second within residues 386-552. Binding of the alpha2 and the gamma1-subunits was not affected by 400 mum AMP or ATP. Furthermore, we show that the beta-subunit C terminus is essential for binding to the alpha2-subunit but, in contrast to previous work, the beta-subunit does not bind directly to the gamma1-subunit. Taken together, this study presents a new model for AMPK heterotrimer structure where through its C terminus the beta-subunit binds to the alpha-subunit that, in turn, binds to the gamma-subunit. There is no direct interaction between the beta- and gamma-subunits.
authors.affiliation: Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA.
authors.forename: Kelly A
authors.initials: KA
authors.lastname: Wong
authors.forename: Harvey F
authors.initials: HF
authors.lastname: Lodish
mesh_terms: D055372:AMP-Activated Protein Kinases; D000255:Adenosine Triphosphate / Q000737:chemistry; D001665:Binding Sites; D015153:Blotting, Western; D020134:Catalytic Domain; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D017353:Gene Deletion; D006801:Humans; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000502:physiology; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000502:physiology; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D014162:Transfection
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D009097:Multienzyme Complexes; D000255:Adenosine Triphosphate; C526181:PRKAA2 protein, human; C526222:PRKAG1 protein, human; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases