K+/H+ antiporter in alkaliphilic Bacillus sp. no. 66 (JCM 9763)

A K+/H+ antiport system was detected for the first time in right-side-out membrane vesicles prepared from alkaliphilic Bacillus sp. no. 66 (JCM 9763). An outwardly directed K+ gradient (intravesicular K+ concentration, K(in), 100 mM; extravesicular K+ concentration, K(out), 0.25 mM) stimulated uphill H+ influx into right-side-out vesicles and created the inside-acidic pH gradient (delta pH). This H+ influx was pH-dependent and increased as the pH increased from 6.8 to 8.4. Addition of 100 microM quinine inhibited the H+ influx by 75%. This exchange process was electroneutral, and the H+ influx was not stimulated by the imposition of the membrane potential (interior negative). Addition of K+ at the point of maximum delta pH caused a rapid K+-dependent H+ efflux consistent with the inward exchange of external K+ for internal H+ by a K+/H+ antiporter. Rb+ and Cs+ could replace K+ but Na+ and Li+ could not. The H+ efflux rate was a hyperbolic function of K+ and increased with increasing extravesicular pH (pH(out)) from 7.5 to 8.5. These findings were consistent with the presence of K+/H+ antiport activity in these membrane vesicles.