Proton motive force during growth of Streptococcus lactis cells

Abstract

Experiments with the aerotolerant anaerobe Streptococcus lactis provide the opportunity for determining the proton motive force (Deltap) in dividing cells. The two components of Deltap, DeltaPsi (the transmembrane potential) and DeltapH (the chemical gradient of H(+)), were determined by the accumulation of radiolabeled tetraphenylphosphonium (TPP(+)) and benzoate ions. The DeltaPsi was calibrated with the K(+) diffusion potential in starved, valinomycin-treated cells. With resting, glycolyzing cells, the Deltap was measured also by the accumulation of the non-metabolizable sugar thiomethyl-beta-galactoside (TMG). In resting cells the Deltap, calculated either by adding DeltaPsi and ZDeltapH or from the levels of TMG, was relatively constant between pH 5 to 7, decreasing from 160 to 150 mV and decreasing further to 100 mV at pH 8.0. With the TPP(+) probe for DeltaPsi, we confirmed our previous finding that the K(+) ions dissipate DeltaPsi and increase DeltapH, whereas Na(+) ions have little effect on DeltaPsi and no effect on DeltapH. [(3)H]TPP(+) and [(14)C]benzoate were added during exponential phase to S. lactis cells growing at pH 5 to 7 at 28 degrees C in a defined medium with glucose as energy source. As with resting cells, the DeltapH and DeltaPsi were dependent on the pH of the medium. At pH 5.1, the DeltapH was equivalent to 60 mV (alkaline inside) and decreased to 25 mV at pH 6.8. The DeltaPsi increased from 83 mV (negative inside) at pH 5.1 to 108 mV at pH 6.8. The Deltap, therefore, was fairly constant between pH 5 and 7, decreasing from 143 to 133 mV. The values for Deltap in growing cells, just as in resting cells, are consistent with a system in which the net efflux of H(+) ions is effected by a membrane-bound adenosine triphosphatase and glycolytically generated adenosine triphosphate. The data suggest that in both growing and resting cells the pH of the medium and its K(+) concentration are the two principal factors that determine the relative contribution of DeltapH and DeltaPsi to the proton motive force.