V-TYPE AND P-TYPE CA2-STIMULATED ATPASES IN A CALCIFYING STRAIN OF PLEUROCHRYSIS SP. (HAPTOPHYCEAE)()

Coccolithophorids are marine unicellular algae characterized by their ability to carry out controlled, subcellular calcification. The bioche mical and kinetic features of membrane-bound Ca2+-stimulated ATPases h ave been examined. Membranes and organelles from axenic cultures of Pl eurochrysis sp. (CCMP299) were isolated by means of sucrose density ce ntrifugation. High levels of Ca2+-stimulated ATPase were detected in c hloroplasts, Golgi apparatus, plasma membrane, and coccolith vesicles. The sensitivity of the enzyme activity in the organelles and membrane s was assessed with pharmacologic agents that are known to be specific for the several isoforms of Ca2+-stimulated ATPase. The Ca2+-stimulat ed ATPase activity in the Golgi and coccolith vesicle preparations was sensitive to nitrate, thiocyanate, and sodium azide and insensitive t o vanadate, cyclopiazonic acid, and thapsigargin. ATP-dependent H+ mov ement, but not Ca-45(2+) transport, across the coccolith vesicle was d emonstrated. The Ca2+-stimulated ATPase in the plasma membrane prepara tion was sensitive to vanadate. ATP-dependent, vanadate-sensitive effl ux of Ca-45(2+) was demonstrated for microsomal material derived from gradient-isolated plasma membrane. Polypeptides from isolated Golgi an d coccolith vesicle preparations cross-reacted to an antibody raised a gainst a subunit of the oat root proton pump, whereas polypeptides fro m the chloroplast preparations did not cross-react. These findings sho w that a V-type Ca2+-stimulated ATPase is located on the coccolith ves icle membrane and a P-type Ca2+-stimulated ATPase is located on the pl asma membrane.