Kinetics of Ca²⁺/H⁺ Antiport in Isolated Tonoplast Vesicles from Storage Tissue of Beta vulgaris L.

Artificial pH gradients across tonoplast vesicles isolated from storage tissue of red beet (Beta vulgaris L.) were used to study the kinetics of a Ca(2+)/H(+) antiport across this membrane. Ca(2+)-dependent H(+) fluxes were measured by the pH-dependent fluorescence quenching of acridine orange. DeltapH-dependent Ca(2+) influx was measured radiometrically. Both H(+) efflux and Ca(2+) influx displayed saturation kinetics and an identical dependence on external calcium with apparent K(m) values of 43.9 and 41.7 micromolar, respectively. Calcium influx was unaffected by an excess of Mg(2+) but was inhibited by La(3+) > Mn(2+) > Cd(2+). The apparent K(m) for external calcium was greatly affected (5-fold) by internal pH in the range of 6.0 to 6.5 and a transmembrane effect of internal proton binding on the affinity for external calcium is suggested.