Yeast protein kinase Ptk2 localizes at the plasma membrane and phosphorylates in vitro the C-terminal peptide of the H+-ATPase.
Glucose triggers posttranslational modifications that increase the activity of the Saccharomyces cerevisiae plasma membrane H+-ATPase (Pma1). Glucose activation of yeast H+-ATPase results from the change in two kinetic parameters: an increase in the affinity of the enzyme for ATP, depending on Ser899, and an increase in the Vmax involving Thr912. Our previous studies suggested that Ptk2 mediates the Ser899-dependent part of the activation. In this study we find that Ptk2 localized to the plasma membrane in a Triton X-100 insoluble fraction. In vitro phosphorylation assays using a recombinant GST-fusion protein comprising 30 C-terminal amino acids of Pma1 suggest that Ser899 is phosphorylated by Ptk2. Furthermore, we show that the Ptk2 carboxyl terminus is essential for glucose-dependent Pma1 activation and for the phosphorylation of Ser899.
Eraso
Pilar
P
Departamento de Bioquímica, Facultad de Medicina, Universidad Autónoma de Madrid and Instituto de Investigaciones Biomédicas Alberto Sols, Consejo Superior de Investigaciones Científicas, Arturo Duperier, 4, 28029 Madrid, Spain.
Mazón
María J
MJ
Portillo
Francisco
F
D000595:Amino Acid Sequence; D001665:Binding Sites; D002462:Cell Membrane / Q000201:enzymology; D004789:Enzyme Activation; D005800:Genes, Fungal; D005947:Glucose / Q000378:metabolism; D007700:Kinetics; D009154:Mutation; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D011505:Protein-Tyrosine Kinases; C092477:PMA1 protein, S cerevisiae; D006180:Proton-Translocating ATPases; D005947:Glucose