Localization of a K+ -binding site involved in dephosphorylation of the sarcoplasmic reticulum Ca2+ -ATPase.
K+ plays an important role for the function of the sarco(endo)plasmic reticulum Ca2+ -ATPase (SERCA), but its binding site within the molecule has remained unidentified. We have located the binding site for a K+ ion in the P-domain by means of x-ray crystallography using crystals prepared in the presence of the K+ congener Rb+. Backbone carbonyls from the loop containing residues 711-715 together with the side chain of Glu732 define the K+/Rb+ site in the Ca2+ -ATPase conformation with bound Ca2+, ADP, and AlF4-. Functional analysis of Ca2+ -ATPase mutants with alterations to Glu732 shows that this site is indeed important for the stimulatory effect of K+ on the dephosphorylation rate. Comparison with the Ca2+ -ATPase in a dephosphorylated E2 conformation suggests that the K+ site is involved in the correct movement and positioning of the A-domain during translocation and dephosphorylation.
Sørensen
Thomas Lykke-Møller
TL
Centre for Structural Biology, Department of Molecular Biology, University of Aarhus, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.
Clausen
Johannes D
JD
Jensen
Anne-Marie Lund
AM
Vilsen
Bente
B
Møller
Jesper Vuust
JV
Andersen
Jens Peter
JP
Nissen
Poul
P
D000244:Adenosine Diphosphate / Q000737:chemistry; D000251:Adenosine Triphosphatases / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D018360:Crystallography, X-Ray; D003593:Cytoplasm / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004583:Electrons; D018698:Glutamic Acid / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010766:Phosphorylation; D011188:Potassium / Q000737:chemistry*; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D013997:Time Factors
D018076:DNA, Complementary; D018698:Glutamic Acid; D000244:Adenosine Diphosphate; D000251:Adenosine Triphosphatases; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D011188:Potassium