SUMMARY: Ca was accumulated by right-side-out membrane vesicles of following imposition of a diffusion potential, inside-negative, owing to K-efflux via valinomycin. Uptake was dependent on the magnitude of the membrane potential. This voltage-dependent Ca uptake was inhibited by Ca channel blockers such as nitrendipine, verapamil and LaCl, and was competitively inhibited by Ba and Sr. The system showed saturation kinetics with an apparent for Ca of about 250 μM. Proteins responsible for the voltage-dependent Ca uptake were partially purified by preparative isoelectric focusing in a Sepharose bed. A fraction at pH 5·28-5·33 contained the activity. The characteristics of Ca uptake in reconstituted proteoliposomes were the same as those in membrane vesicles (sensitive to Ca channel blockers; inhibited by Ba and Sr). In addition, uptake was not influenced by a pH gradient imposed on the vesicles. The apparent for Ca in the reconstituted system was about 260 μM. The specific activity was increased about 50-fold by purification with isoelectric focusing.


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