Calcium transport and Ca2+-ATPase activity in ram spermatozoa plasma membrane vesicles

Plasma membrane vesicles, isolated from ejaculated ram sperm, were found to contain Ca2+-activated Mg2+-ATPase and Ca2+ transport activities. Membrane vesicles that were exposed to oxalate as a Ca2+-trapping agent accumulated Ca2+ in the presence of Mg2+ and ATP. The Vmax for Ca2+ uptake was 33 nmol/mg protein per h, and the Km values for Ca2+ and ATP were 2.5 microM and 45 microM, respectively. 1 microM of the Ca2+ ionophore A23187, added initially, completely inhibited net Ca2+ uptake and, if added later, caused the release of Ca2+ previously accumulated. A Ca2+-activated ATPase was present in the same membrane vesicles which had a Vmax of 1.5 mumol/mg protein per h at free Ca2+ concentration of 10 microM. This Ca2+-ATPase had Km values of 4.5 microM and 110 microM for Ca2+ and ATP, respectively. This kinetic parameter was similar to that observed for uptake of Ca2+ by the vesicles. The Ca2+-ATPase activity was insensitive to ouabain. Both Ca2+ transport and Ca2+-ATPase activity were inhibited by the flavonoid quercetin. Thus, ram spermatozoa plasma membranes have both a Ca2+ transport activity and a Ca2+-stimulated ATPase activity with similar substrate affinities and specificities and similar sensitivity to quercetin.