Abstract
Antibodies raised in rabbits against the purified erythrocyte membrane Ca2+ pumping ATPase were affinity-purified using an ATPase-Sepharose column. Addition of a few molecules of the purified antibody per molecule of ATPase was sufficient to inhibit the ATPase activity. Extensively washed ghosts or preincubated pure ATPase sometimes develop an appreciable Mg2+-ATPase activity. In such cases, the antibodies inhibited the Mg2+-ATPase as well as the Ca2+-ATPase. This is consistent with the hypothesis that a portion of the Mg2+-ATPase activity of ghosts is derived from the Ca2+-ATPase. When nitrophenylphosphatase activity was observed, both Mg2+ - and Ca2+-stimulated activities were observed. Only the Ca2+ activity was inhibited by the antibodies, confirming that this activity is due to the Ca2+ pump, and suggesting that the Mg2+-nitrophenylphosphatase is due to a separate enzyme. Amounts of antibody comparable to those which inhibited the Ca2+-ATPases had no effect on the Na+-K+-ATPase; 4-fold higher amounts of antibody significantly stimulated the Na+-K+-ATPase, but this effect of the antibody was not specific: Immunoglobulins from the nonimmune serum also significantly stimulated the Na+-K+-ATPase.
In resealed erythrocyte membranes, antibodies incorporated into the ghosts inactivated the Ca2+-ATPase, while antibodies added to the outside had no significant effect.
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Verma, A.K., Penniston, J.T., Muallem, S. et al. Effects of affinity-purified antibodies on the Ca2+ pumping ATPase of erythrocyte membranes. J Bioenerg Biomembr 16, 365–378 (1984). https://doi.org/10.1007/BF00743232
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DOI: https://doi.org/10.1007/BF00743232