A conserved sequence immediately N-terminal to the Bateman domains in AMP-activated protein kinase gamma subunits is required for the interaction with the beta subunits.
Mammalian AMP-activated protein kinase is a serine/threonine protein kinase that acts as a sensor of cellular energy status. AMP-activated protein kinase is a heterotrimer of three different subunits, i.e. alpha, beta, and gamma, with alpha being the catalytic subunit and beta and gamma having regulatory roles. Although several studies have defined different domains in alpha and beta involved in the interaction with the other subunits of the complex, little is known about the regions of the gamma subunits involved in these interactions. To study this, we have made sequential deletions from the N termini of the gamma subunit isoforms and studied the interactions with alpha and beta subunits, both by two-hybrid analysis and by co-immunoprecipitation. Our results suggest that a conserved region of 20-25 amino acids in gamma1, gamma2, and gamma3, immediately N-terminal to the Bateman domains, is required for the formation of a functional, active alphabetagamma complex. This region is required for the interaction with the beta subunits. The interaction between the alpha and gamma subunits does not require this region and occurs instead within the Bateman domains of the gamma subunit, although the alpha-gamma interaction does appear to stabilize the beta-gamma interaction. In addition, sequential deletions from the C termini of the gamma subunits indicate that deletion of any of the CBS (cystathionine beta-synthase) motifs prevents the formation of a functional complex with the alpha and beta subunits.
Viana
Rosa
R
Instituto de Biomedicina de Valencia, Consejo Superior de Investigaciones Científicas, Jaime Roig 11, 46010 Valencia, Spain.
Towler
Mhairi C
MC
Pan
David A
DA
Carling
David
D
Viollet
Benoit
B
Hardie
D Grahame
DG
Sanz
Pascual
P
D055372:AMP-Activated Protein Kinases; D020816:Amino Acid Motifs / Q000235:genetics; D020134:Catalytic Domain / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D009097:Multienzyme Complexes; D011485:Protein Binding / Q000235:genetics; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D017434:Protein Structure, Tertiary / Q000235:genetics; D017384:Sequence Deletion / Q000235:genetics; D020798:Two-Hybrid System Techniques
D009097:Multienzyme Complexes; D020033:Protein Isoforms; D011494:Protein Kinases; C526222:PRKAG1 protein, human; C526225:PRKAG3 protein, human; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases