Generation of an electrochemical proton gradient by lactate efflux in membrane vesicles of Escherichia coli.

The 'energy-recycling model' [Michels et al. (1979) FEMS Microbiol. Lett. 5, 357-364] postulates the generation of an electrochemical gradient across the bacterial cytoplasmic membrane by carrier-mediated efflux of metabolic endproducts in symport with protons. Experimental evidence for this model is presented. In membrane vesicles from Escherichia coli ML 308-255 L-lactate translocation (both uptake and efflux) is carrier-mediated. The H+/L-lactate stoichiometry varies, depending on the external pH, between 1 and 2. This change in stoichiometry is most likely the result of a protonation of the lactate carrier protein. This process has a pK of 6.75. L-Lactate efflux from membrane vesicles, loaded with 50 mM potassium L-lactate, results at an external pH of 6.6 in an 11-fold accumulation of proline inside the vesicles. This accumulation is completely inhibited by the uncoupler carbonyl cyanide p-trifluoromethoxyphenylhydrazone. The uptake of proline is not the result of a potassium or an osmotic gradient. At an external pH of 6.6 efflux of L-lactate from the vesicles leads to the generation of an electrical potential across the membrane of -55 mV, as is demonstrated from the accumulation of the lipophilic cation tetraphenylphosphonium.