Phosphorylation of the Na,K-ATPase alpha-subunit by protein kinase A and C in vitro and in intact cells. Identification of a novel motif for PKC-mediated phosphorylation.

Détails

ID Serval
serval:BIB_4973DA3E2CA8
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Titre
Phosphorylation of the Na,K-ATPase alpha-subunit by protein kinase A and C in vitro and in intact cells. Identification of a novel motif for PKC-mediated phosphorylation.
Périodique
Journal of Biological Chemistry
Auteur(s)
Beguin P., Beggah A.T., Chibalin A.V., Burgener-Kairuz P., Jaisser F., Mathews P.M., Rossier B.C., Cotecchia S., Geering K.
ISSN
0021-9258 (Print)
ISSN-L
0021-9258
Statut éditorial
Publié
Date de publication
1994
Volume
269
Numéro
39
Pages
24437-24445
Langue
anglais
Résumé
Na,K-ATPase is a potential target for regulatory phosphorylation by protein kinase A and C (PKA and PKC). To identify the phosphorylation sites, we have mutated the alpha 1-subunit of Bufo marinus in a highly conservative PKA and in 20 different PKC consensus sequences. The mutants were expressed in Xenopus oocytes and their phosphorylation capacity tested in homogenates upon stimulation of PKA or PKC. While serine 943 (Ser-943) was identified as a unique target site for PKA, none of the PKC consensus serine or threonine residues are implicated in PKC phosphorylation. Controlled trypsinolysis of phosphorylated alpha-subunits of various purified enzyme preparations and of alpha/beta complexes from oocyte homogenates revealed that PKC phosphorylation was exclusively associated with the N terminus. A fusion protein containing the first 32 amino acids of the Bufo alpha-subunit was phosphorylated in vitro and serine and threonine residues (Thr-15 and Ser-16) in this region were identified by site-directed mutagenesis as the PKC phosphorylation sites. Finally, the Bufo alpha-subunit was phosphorylated by protein kinases in transfected COS-7 cells. In intact cells, PKA stimulation induced phosphorylation exclusively on Ser-943 and PKC stimulation mainly on Thr-15 and Ser-16, which are contained in a novel PKC phosphorylation motif.
Mots-clé
Amino Acid Sequence, Animals, Base Sequence, Bufo marinus, Cells, Cultured, Cyclic AMP-Dependent Protein Kinases/metabolism, Molecular Sequence Data, Mutagenesis, Site-Directed, Oligodeoxyribonucleotides, Oocytes, Phosphorylation, Protein Kinase C/metabolism, Sodium-Potassium-Exchanging ATPase/chemistry, Sodium-Potassium-Exchanging ATPase/metabolism, Xenopus laevis
Pubmed
Web of science
Création de la notice
24/01/2008 12:05
Dernière modification de la notice
03/03/2018 16:54
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