| A marine unicellular alga, the coccolithophorid Pleurochrysis CCMP-299, contains specialized organelles, in which CaCO{dollar}sb3{dollar} crystals are formed on a baseplate to complete a coccolith. Coccoliths are exocytosed and form a coccosphere around the cell. For this intracellular calcification, protons are thought to be transported actively out of the coccolith vesicles. This study investigated the Ca{dollar}sp{lcub}2+{rcub}{dollar}-stimulated ATPase activity at the organelle level. Coccolith and Golgi vesicles demonstrated Ca{dollar}sp{lcub}2+{rcub}{dollar}-stimulated ATPase activities which were sensitive to V-type ATPase inhibitors. Their 60 kDa polypepetides cross-reacted to an anti-V-type-H{dollar}sp+{dollar}-ATPase subunit from oat. A significant ATP-dependent outward movement of protons was also detected on the coccolith-vesicle membrane. From the studies of the localization, kinetic, and biochemical properties of the Ca{dollar}sp{lcub}2+{rcub}{dollar}-stimulated ATPase and H{dollar}sp+{dollar}-pump, it is concluded that H{dollar}sp+{dollar}-pump activity due to the Ca{dollar}sp{lcub}2+{rcub}{dollar}-stimulated ATPase is present on the coccolith-vesicle membrane. Experiments with {dollar}rmsp{lcub}45{rcub}Casp{lcub}2+{rcub}{dollar} showed no ATP-dependent Ca{dollar}sp{lcub}2+{rcub}{dollar}-transport activity on the coccolith-vesicle membrane, indicating that the enzyme is not a {dollar}rm Casp{lcub}2+{rcub}/Hsp+{dollar}-antiporter. However, Ca{dollar}sp{lcub}2+{rcub}{dollar}-binding activity, presumably the property of Ca{dollar}sp{lcub}2+{rcub}{dollar}-binding polysaccharides, was significant on the coccolith vesicles.; The plasma membrane also demonstrated Ca{dollar}sp{lcub}2+{rcub}{dollar}-stimulated ATPase activity, but this enzyme was found to be of the P-type. There was an ATP-dependent outward Ca{dollar}sp{lcub}2+{rcub}{dollar}-transport activity on the plasma membrane suggesting a Ca{dollar}sp{lcub}2+{rcub}{dollar}-eliminating activity even in the calcifying coccolithophorid. Ca{dollar}sp{lcub}2+{rcub}{dollar}-binding polysaccharides were also found on the plasma membrane. |
