A novel primary Ca(2+)-transport system from Saccharomyces cerevisiae

A novel primary Ca(2+)-transport system in membranes from Saccharomyces cerevisiae is described. Ca2+ transport is strictly dependent on the presence of ATP; other nucleotides like GTP, UTP and CTP do not efficiently (< 10% of the rate of ATP) drive uptake. Transport is inhibited by sodium vanadate with an IC50 of 130 microM, but is insensitive to carbonylcyanide p-trifluoromethoxy-phenylhydrazone, valinomycin, gramicidin or calmodulin. Ca2+ accumulates in a free form and can be readily released by the Ca2+ ionophore A-23187 or by osmotic shock. The apparent Km values of transport activity for free Ca2+ was determined to be 0.11 microM and 5 microM for Mg.ATP, respectively. Taken together the results indicate that the Ca2+ transport described here does not belong to the plasma-membrane-type Ca(2+)-ATPase family but rather to the family of endomembrane-type ATPases. Cell-fractionation studies of crude membranes on sucrose gradient centrifugation have shown that the Ca(2+)-transport activity separates from marker enzymes for endoplasmic reticulum, vacuole, or plasma membrane and migrates with GDPase activity, a marker for the yeast Golgi complex.