Relationships between the sodium-proton antiport activity and the components of the electrochemical proton gradient in Escherichia coli membrane vesicles

Abstract

The kinetics of Na+ efflux from E. coli RA 11 membrane vesicles taking place along a favorable Na+ concentration gradient are strongly dependent on the generation of an electrochemical proton gradient. An energy-dependent acceleration of the Na+ efflux rate is observed at all external pH of 5.5-7.5 and is prevented by uncoupling agents. The contributions of the electrical potential (.DELTA..psi.) and chemical potential (.DELTA.pH) of H+ to the mechanism of Na+ efflux acceleration were studied by determining the effects of selective dissipation of .DELTA..psi. and .DELTA.pH in respiring membrane vesicles with valinomycin or nigericin and imposition of outwardly directed K+ diffusion gradients (imposed .DELTA..psi., interior negative) or acetate diffusion gradients (imposed .DELTA.pH, interior alkaline). At pH 6.6 and 7.5, .DELTA.pH and .DELTA..psi. individually and concurrently accelerate the downhill Na+ efflux rate. At pH 5.5, the Na+ efflux rate is enhanced by .DELTA.pH only when the imposed .DELTA.pH exceeds a threshold .DELTA.pH value; an imposed .DELTA..psi. which per se does not enhance the Na+ efflux rate does not contribute to the acceleration of Na+ efflux when imposed simultaneously with a .DELTA.pH higher than the threshold .DELTA.pH value. The results strongly suggest that the Na+-H+ antiport mechanism catalyzes the downhill Na+ efflux and suggest that the overall exchange reaction is rate limited by the rate of coupled H+ influx. The Na+-H+ antiporter might function as an electrogenic process at all pH of 5.5-7.5 and the antiport function is controlled, in particular at acidic pH, by a .DELTA.pH-sensitive reaction or alternatively by the internal pH.