Interaction between poly(L-lysine) and membranes inhibits proton pumping by corn root tonoplast H+-ATPase

The influence of poly(L-lysine) binding on the coupled activities of nitrate-sensitive H⁺-ATPase in isolated corn ( Zea mays L. cv. FRB73) root tonoplast vesicles was investigated. The addition of membrane-impermeable poly(L-lysine) caused a slow increase in light scattering of the tonoplast suspension. Electron microscopy showed that the increase was the result of an aggregation of the vesicles. In the presence of 75 m M KCl, a concentration sufficient to sustain near optimal ATP hydrolysis, poly(L-lysine) slightly enhanced the hydrolysis activity but significantly inhibited proton pumping of the H⁺-ATPase. Inhibition increased with the average molecular mass of poly(L-lysine) and reached a maximum at 58 kDa. When total osmolarity was kept constant, the replacement of sucrose by KCl enhanced both ATP hydrolysis and proton pumping activities. However, enhancement of proton pumping was significantly greater than that of ATP hydrolysis. An increase in KCl, but not K₂SO₄, significantly relieved poly(L-lysine)-induced inhibition of proton pumping. Kinetic analysis indicated that poly(L-lysine) did not significantly affect the proton leakage of the tonoplast membranes under different energetic conditions. These results suggest that the electrostatic interaction between poly(L-lysine) and the negative charges on the exterior surface of tonoplast vesicles could change the coupling ratio of ATP hydrolysis to proton pumping. Thus, the surface charge of the tonoplast membrane may be involved in the regulation of these two activities.