Kinetics of calcium transport through purified plasma membrane of buffalo (Bubalus bubalis) spermatozoa: effect of calmodulin-like protein.

Abstract

A procedure is described for the isolation, identification, and purification of plasma membrane from buffalo ejaculated spermatozoa. A typical yield of plasma membrane vesicles obtained by this procedure was 280 micrograms of membrane protein per (10)9 spermatozoa. Kinetics of calcium transport through purified plasma membrane vesicles were studied using 45Ca2+. The transport of Ca2+ through plasma membrane vesicles was electrogenic with maximum Ca2+ uptake (47 mumol) occurring at 30 min in the presence of 0.25 mM ATP. Km and V(max) values for Ca2+ transport were 0.25 mM and 182 mumol, respectively, whereas in the presence of variable concentrations of free Ca2+, the Km and V(max) values were 0.23 mumol and 82.3 mumol, respectively. Calmodulin-like protein (20 micrograms) significantly (p < 0.01) inhibited the Ca2+ accumulation in the plasma membrane vesicles. It is demonstrated that calmodulin-like protein regulates Ca2+ transport during sperm capacitation and acrosome reaction.