THE VOLTAGE-DEPENDENT H-ATPASE OF THE SUGAR-BEET VACUOLE IS REVERSIBLE()

The vacuolar H+-ATPase is essential for the creation and maintenance o f solute gradients. Knowledge of the reversal potential, expressed by the voltage and pH dependence of the pump may allow to determine the a ctivity range of the enzyme. In the whole-vacuole configuration of the patch-clamp technique the application of Mg-ATP elicited inward-direc ted currents through the H+-ATPase. Reversal of the pump current was o btained in the presence of a pH gradient across the membrane (inside a cid) by replacement of Mg-ATP by Mg-ADP and P-i. The active nature of this nucleotide-dependent transport process is reflected by a Q(10) of 3.2. The voltage-dependence of the pump was elucidated by voltage-ste ps to various depolarizing and hyperpolarizing potentials. In the pres ence of Mg-ATP the current-voltage relationship of the pump current is characterized by an almost linear increase of the steady state curren t between 20 mV and 100 mV, tending to saturate at more positive poten tials. The voltage-dependence of the inward pump current could be desc ribed by a pseudo-two-state model.