Symport of proton and sucrose in plasma membrane vesicles isolated from spinach leaves

The mechanism of sucrose transport was investigated in plasma membrane (PM) vesicles isolated from spinach (Spinacia oleracea L.) leaves. PM vesicles were isolated by aqueous two-phase partitioning and were equilibrated in pH 7.8 buffer containing K+. The vesicles rapidly accumulated sucrose in the presence of a transmembrane pH gradient (deltapH) with external pH set at 5.8. The uptake rate was slow at pH 7.8. The K+-selective ionophore, valinomycin, stimulated uptake in the presence of a deltapH, and the protonophore, carbonyl cyanide m-chlorophenylhydrazone (CCCP), greatly inhibited deltapH-dependent sucrose uptake. Addition of sucrose to the vesicles resulted in immediate alkalization of the medium. Alkalization was stimulated by valinomycin, was abolished by CCCP, and was sucrose-specific. These results demonstrate the presence of a tightly coupled H+/sucrose symporter in PM vesicles isolated from spinach leaves