Cooperative setting for long-range linkage of Ca(2+) binding and ATP synthesis in the Ca(2+) ATPase.
High-affinity and cooperative binding of two Ca(2+) per ATPase (SERCA) occurs within the membrane-bound region of the enzyme. Direct measurements of binding at various Ca(2+) concentrations demonstrate that site-directed mutations within this region interfere selectively with Ca(2+) occupancy of either one or both binding sites and with the cooperative character of the binding isotherms. A transition associated with high affinity and cooperative binding of the second Ca(2+) and the engagement of N796 and E309 are both required to form a phosphoenzyme intermediate with ATP in the forward direction of the cycle and also to form ATP from phosphoenzyme intermediate and ADP in the reverse direction of the cycle. This transition, defined by equilibrium and kinetic characterization of the partial reactions of the enzyme cycle, extends from transmembrane helices to the catalytic site through a long-range linkage and is the mechanistic device for interconversion of binding and phosphorylation potentials.
Inesi
Giuseppe
G
Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, 108 N. Greene Street, Baltimore, MD 21201, USA. ginesi@umaryland.edu
Zhang
Zhongsen
Z
Lewis
David
D
D000244:Adenosine Diphosphate / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D019556:COS Cells; D002118:Calcium / Q000378:metabolism*; D020134:Catalytic Domain; D002645:Chickens; D004305:Dose-Response Relationship, Drug; D007477:Ions; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology; D008958:Models, Molecular; D009154:Mutation; D010766:Phosphorylation; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors
D007477:Ions; D011994:Recombinant Proteins; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D002118:Calcium