Quantitative Analysis of ATP-Dependent H^+ Efflux and Pump Current Driven by an Electrogenic Pump in Nitellopsis obtusa

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Internodal cells of Nitellopsis were made tonoplast-free by perfusion with a medium containing EGTA. Cytoplasmic concentrations of solutes were controlled by a second perfusion with media of known composition. The electrogenic pump current (I_p), which was calculated from electrical data obtained from cells with and without ATP, was compared with the current carried by H+ (I_H+) across the plasma membrane. A close correlation between I_p and I_H+ was found under various internal and external conditions. (1) I_p and I_H+ depended on the internal ATP. and showed Michaelis-Menten type saturation curves. For I_p, K_m was 120μM and the maximum current V_<max> was 15.1mAm^<-2>.(2) I_p_ and I_H+ showed almost the same Mg<2+> dependence. The Mg^<2+>-dependent I_p was 19.5 mAm^<-2> while the Mg^<2+>-dependent I_H+ was 17.7 mAm^<-2> (3) I_H+ was maximal at an external pH of 8 and decreased both in acidic and alkaline pH ranges. I_p was nearly equal to I_H+ in the pH range between 8 and 5. (4) I_H+ became maximal at an internal pH of 7.3, which is nearly the same as the pH for maximal electrogenecity found by Mimura and Tazawa (1984). All these facts support the idea proposed in our previous paper (Takeshige et al. 1985) that the electrogenic ion pump located in the plasma membrane of Nitellopsis is the H^+ pump.
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