A kinetic analysis of the electrogenic pump ofChara corallina: II. Dependence of the pump activity on external pH

Abstract

The current-voltage curve of theChara membrane was obtained by applying a slow ramp de- and hyperpolarization by use of voltage clamp. By inhibiting the electrogenic pump with 50μm DCCD (dicyclohexylcarbodiimide), theI–V curve approached a steady state within 100 min, which gave thei d -V curve of the passive diffusion channel. Thei p -V curve of the electrogenic pump channel was obtained by subtracting the latter from the former. With the increase of external pH, thei d -V curve showed only a slight change, while thei p -V curve of the pump channel showed almost a parallel shift, in the hyperpolarizing direction, along the voltage axis in the pH range between 6.5 and 7.5. The sigmoidali p -V curve in this pH range could be simulated satisfactorily with the five-state model reported previously (U. Kishimoto, N. Kami-ike, Y. Takeuchi & T. Ohkawa,J. Membrane Biol.80:175–183, 1984) as well as with a lumped two-state model presented in this report. The analysis based on these models suggests that the electrogenic pump of theChara membrane is mainly a 2H+/1ATP pump. The forward rate constant in the voltage-dependent step increased with the increase of external pH, while the backward one decreased. On the other hand, the forward rate constant in the voltage-independent step remained almost unchanged with the increase of external pH, while the backward one increased markedly. The pump conductance at the resting membrane potential showed either a slight increase or a decrease with the increase of external pH, depending on the sample. Nevertheless, the pump current showed generally a slight increase with the increase of external pH.