Proton efflux during Ca2+ uptake in a reconstituted Ca2+ pump system

Ca2+-pump of sarcoplasmic reticulum was partially purified and solubilized with Triton-X100 in the presence of excess phospholipids. Vesicles were reconstituted by removal of Triton with Amberlite XAD-2 and the subsequent freeze-thaw/sonication method (Kasahara, M. and Hinkle, P. C. (1977) J. Biol. Chem., 252: 7384-7390). In the reconstituted system, H+ was ejected during Ca2+ uptake and in the initial Ca2+ uptake, the molar ratio of ejected H+ to Ca2+ was found to be about 1 in the presence of a permeant anion, SCN-.