Structure of the Na,K-ATPase regulatory protein FXYD1 in micelles.
FXYD1 is a major regulatory subunit of the Na,K-ATPase and the principal substrate of hormone-regulated phosphorylation by c-AMP dependent protein kinases A and C in heart and skeletal muscle sarcolemma. It is a member of an evolutionarily conserved family of membrane proteins that regulate the function of the enzyme complex in a tissue-specific and physiological-state-specific manner. Here, we present the three-dimensional structure of FXYD1 determined in micelles by NMR spectroscopy. Structure determination was made possible by measuring residual dipolar couplings in weakly oriented micelle samples of the protein. This allowed us to obtain the relative orientations of the helical segments and information about the protein dynamics. The structural analysis was further facilitated by the inclusion of distance restraints, obtained from paramagnetic spin label relaxation enhancements, and by refinement with a micelle depth restraint, derived from paramagnetic Mn line broadening effects. The structure of FXYD1 provides the foundation for understanding its intra-membrane association with the Na,K-ATPase alpha subunit and suggests a mechanism whereby the phosphorylation of conserved Ser residues, by protein kinases A and C, could induce a conformational change in the cytoplasmic domain of the protein to modulate its interaction with the alpha subunit.
Teriete
Peter
P
Burnham Institute for Medical Research, 10901 North Torrey Pines Road, La Jolla, California 92037, USA.
Franzin
Carla M
CM
Choi
Jungyuen
J
Marassi
Francesca M
FM
D000818:Animals; D045744:Cell Line, Tumor; D004578:Electron Spin Resonance Spectroscopy; D005260:Female; D005612:Freeze Drying; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D008823:Micelles; D010750:Phosphoproteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D014981:Xenopus
D008565:Membrane Proteins; D008823:Micelles; D010750:Phosphoproteins; D011994:Recombinant Proteins; C069141:phospholemman; D000254:Sodium-Potassium-Exchanging ATPase