Effects of PKA phosphorylation on the conformation of the Na,K-ATPase regulatory protein FXYD1.
FXYD1 (phospholemman) is a member of an evolutionarily conserved family of membrane proteins that regulate the function of the Na,K-ATPase enzyme complex in specific tissues and specific physiological states. In heart and skeletal muscle sarcolemma, FXYD1 is also the principal substrate of hormone-regulated phosphorylation by c-AMP dependent protein kinase A and by protein kinase C, which phosphorylate the protein at conserved Ser residues in its cytoplasmic domain, altering its Na,K-ATPase regulatory activity. FXYD1 adopts an L-shaped alpha-helical structure with the transmembrane helix loosely connected to a cytoplasmic amphipathic helix that rests on the membrane surface. In this paper we describe NMR experiments showing that neither PKA phosphorylation at Ser68 nor the physiologically relevant phosphorylation mimicking mutation Ser68Asp induces major changes in the protein conformation. The results, viewed in light of a model of FXYD1 associated with the Na,K-ATPase alpha and beta subunits, indicate that the effects of phosphorylation on the Na,K-ATPase regulatory activity of FXYD1 could be due primarily to changes in electrostatic potential near the membrane surface and near the Na(+)/K(+) ion binding site of the Na,K-ATPase alpha subunit.
Teriete
Peter
P
Burnham Institute for Medical Research, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA.
Thai
Khang
K
Choi
Jungyuen
J
Marassi
Francesca M
FM
D000595:Amino Acid Sequence; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011487:Protein Conformation; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
D008565:Membrane Proteins; D010750:Phosphoproteins; C069141:phospholemman; D017868:Cyclic AMP-Dependent Protein Kinases; D000254:Sodium-Potassium-Exchanging ATPase