ENERGY TRANSDUCTION IN THE THERMOPHILIC ANAEROBIC BACTERIUM CLOSTRIDIUM-FERVIDUS IS EXCLUSIVELY COUPLED TO SODIUM-IONS

The thermophilic, peptidolytic, anaerobic bacterium Clostridium fervidus is unable to generate a pH gradient in the range of 5.5-8.0, which limits growth of the organism to a narrow pH range (6.3-7.7). A significant membrane potential (DELTApsi almost-equal-to -60 mV) and chemical gradient of Na+ (-ZDELTApNa almost-equal-to -60 mV) are formed in the presence of metabolizable substrates. Energy-dependent Na+ efflux is inhibited by the Na+/H+ ionophore monensin but is stimulated by uncouplers, suggesting that the Na+ gradient is formed by a primary pumping mechanism rather than by secondary Na+/H+ antiport. This primary sodium pump was found to be an ATPase that has been characterized in inside-out membrane vesicles and in proteoliposomes in which solubilized ATPase was reconstituted. The enzyme is stimulated by Na+, resistant to vanadate, and sensitive to nitrate, which is indicative of an F/V-type Na+-ATPase. In the proteoliposomes Na+ accumulation depends on the presence of ATP, is inhibited by the ATPase inhibitor nitrate, and is completely prevented by the ionophore monensin but is stimulated by protonophores and valinomycin. These and previous observations, which indicated that secondary amino acid transport uses solely Na+ as coupling ion, demonstrate that energy transduction at the membrane in C. fervidus is exclusively dependent on a Na+ cycle.