Cross-phosphorylation between Arabidopsis thaliana sucrose nonfermenting 1-related protein kinase 1 (AtSnRK1) and its activating kinase (AtSnAK) determines their catalytic activities.
Arabidopsis thaliana sucrose nonfermenting 1-related protein kinase 1 complexes belong to the SNF1/AMPK/SnRK1 protein kinase family that shares an ancestral function as central regulators of metabolism. In A. thaliana, the products of AtSnAK1 and AtSnAK2, orthologous to yeast genes, have been shown to autophosphorylate and to phosphorylate/activate the AtSnRK1.1 catalytic subunit on Thr(175). The phosphorylation of these kinases has been investigated by site-directed mutagenesis and tandem mass spectrometry. The autophosphorylation site of AtSnAK2 was identified as Thr(154), and it was shown to be required for AtSnAK catalytic activity. Interestingly, activated AtSnRK1 exerted a negative feedback phosphorylation on AtSnAK2 at Ser(261) (Ser(260) of AtSnAK1) that was dependent on AtSnAK autophosphorylation. The dynamics of these reciprocal phosphorylation events on the different kinases was established, and structural modeling allowed clarification of the topography of the AtSnAK phosphorylation sites. A mechanism is proposed to explain the observed changes in the enzymatic properties of each kinase triggered by these phosphorylation events.
Crozet
Pierre
P
From the Laboratoire de Signalisation et Régulation Métabolique, Institut de Biologie des Plantes, Université Paris-Sud 11, UMR 8618 CNRS, Bâtiment 630, 91405 Orsay Cedex, France.
Jammes
Fabien
F
Valot
Benoit
B
Ambard-Bretteville
Françoise
F
Nessler
Sylvie
S
Hodges
Michael
M
Vidal
Jean
J
Thomas
Martine
M
D019943:Amino Acid Substitution; D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D004789:Enzyme Activation; D025461:Feedback, Physiological; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011487:Protein Conformation; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013395:Sucrose / Q000378:metabolism; D053719:Tandem Mass Spectrometry
D029681:Arabidopsis Proteins; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D021122:Protein Subunits; D011994:Recombinant Proteins; D013395:Sucrose; C531469:At3g01090 protein, Arabidopsis