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Ca 2+ ‐dependent ATPase associated with plasma membrane from a calcareous alga, Serraticardia maxima (Corallinaceae, Rhodophyta)
Author(s) -
Mori Izumi C.,
Sato Gorou,
Okazaki Megumi
Publication year - 1996
Publication title -
phycological research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.438
H-Index - 44
eISSN - 1440-1835
pISSN - 1322-0829
DOI - 10.1111/j.1440-1835.1996.tb00049.x
Subject(s) - vesicle , atpase , nigericin , biology , biochemistry , vanadate , membrane , oligomycin , enzyme
SUMMARY The plasma membrane was isolated from a calcareous red alga, Serraticardia maxima (Yendo) Silva (Corallinaceae), by aqueous two‐phase partitioning. Its purity was examined with marker enzymes, Mg 2+ ‐dependent ATPase, inosine diphosphatase, cytochrome c oxidase and NADH‐cytochrome c reductase, as well as the sensitivity of Mg 2+ ‐dependent ATPase to vanadate, azide and nitrate. The results showed that the isolated plasma membrane was purified enough to study its functions. Electron microscopic observations on thin tissue sections revealed that most vesicles of the isolated plasma membrane were stained by the plasma membrane specific stain, phosphotungstic acid‐chromic acid. Mg 2+ ‐ or Ca 2+ ‐dependent ATPases were associated with the plasma membrane. Ca 2+ ‐dependent ATPase was activated at physiological cytoplasmic concentrations of Ca 2+ (0.1–10 μmol/L). However, calmodulin (0.5 μmol/L) did not affect its activity. The pH optimum was 8.0, in contrast to 7.0 for Mg 2+ ‐dependent ATPase. The isolated plasma membrane vesicles were mostly right side‐out. To test for H + ‐translocation, right side‐out vesicles were inverted; 27% of vesicles were inside‐out after treatment with Triton X‐100. The inside‐out plasma membrane vesicles showed reduction of quinacrine fluorescence in the presence of 1 mmol/L ATP and 100 μmol/L Ca 2+ . The reduced fluorescence was recovered with the addition of 10 mmol/L NH 4 Cl, or 5 μmol/L nigericin plus 50 mmol/L KCl. UTP and CTP substituted for ATP, but ADP did not. Ca 2+ ‐dependent ATPase might pump H + out in the physiological state. The acidification by this pump might be coupled with alkalinization at the calcifying sites, which induces calcification.