The Effects of Cations on Ouabain Binding by Human Erythrocytes
Rodgers, John C.
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Ouabain binding by the human erythrosyte membrane is reversible, exhibits a high of chemical specificity, and can be detected at ouabain concentrations as low as lxlO-l0M. The relation between ouabain binding and ouabain concentration can be described by a rectangular hyperbola permitting determination of the maximal, binding (Bmax) and the ouabain concentration at which ouabain binding is half-maximal (KB). In the presence of magnesium (O.4mM) there was a significant increase of ouabain binding to solubilized erthrosyte membranes and Bmax was reached at 10-5 M ATP. There was stimulation with both magnesium and ATP or with magnesium alone, but stimulation of ouabain was three times greater in the presence of ATP. Oalcium and barium were used in place of magnesium. Calcium, but not barium, produced similar effects to those of magnesium. Other monovalent cations inhibited ouabain binding in the presence of ATP and showed little stimulation in the absence of ATP. Ouabain binding was abolished by proteolytic enzymes and by the sulfhydral agent POMB. This suggests that ouabain probably binds to a protein. Nucleotides other than ATP were studied, and all stimulated ouabain binding both in the presence and absence of magnesium.