Respiratory-driven Na+ electrical potential in the bacterium Vitreoscilla

Vitreoscilla is a Gram-negative bacterium with unique respiratory physiology in which Na+ was implicated as a coupling cation for the generation of a transmembrane electrical gradient (delta psi). Thus, cells respiring in the presence of 110 mM Na+ generated a delta psi of -142 mV compared to only -42 and -56 mV for Li+ and choline, respectively, and even the -42 and -56 mV were insensitive to the protonophore 3,5-di-tert-butyl-4-hydroxybenzaldehyde (DTHB). The kinetics of delta psi formation and collapse correlated well with the kinetics of Na+ fluxes but not with those of H+ fluxes. Cyanide inhibited respiration, Na+ extrusion, and delta psi formation 81% or more, indicating that delta psi formation and Na+ extrusion were coupled to respiration. Experiments were performed to distinguish among three possible transport systems for this coupling: (1) a Na(+)-transporting ATPase; (2) an electrogenic Na+/H+ antiport system; (3) a primary Na+ pump directly driven by the free energy of electron transport. DCCD and arsenate decreased cellular ATP up to 86% but had no effect on delta psi, evidence against a Na(+)-transporting ATPase. Low concentrations of DTHB had no effect on delta psi; high concentrations transiently collapsed delta psi, but led to a stimulation of Na+ extrusion, the opposite of that expected for a Na+/H+ antiport system. Potassium ion, which collapses delta psi, also stimulated Na+ extrusion. The experimental evidence is against Na+ extrusion by mechanisms 1 and 2 and supports the existence of a respiratory-driven primary Na+ pump for generating delta psi in Vitreoscilla.