Coupled Transport of Citrate and Magnesium in Bacillus subtilis

Citrate transport in Bacillus subtilis is dependent on the presence of Mg2+. Other divalent cations like Mn2+, Co2+, or Ni2+ can replace magnesium ions. 28Mg2+ or 54Mn2+ are taken up by the cells simultaneously with the uptake of citrate. Cells which are not induced for citrate transport do not show the citrate-dependent uptake of either 54Mn2+ or 28Mg2+. Citrate transport and the citrate-dependent transport of a divalent cation are inhibited by the uncoupling agent m-chlorophenyl carbonylcyanidehydrazone. Therefore, using this uncoupler, it is possible to distinguish between the citrate-independent binding of divalent cations to the cell surface and the citrate-dependent transport of these ions across the cytoplasmic membrane. The kinetic constants of citrate uptake and of citrate-dependent Mg2+ transport are very similar. Citrate transport may be stoichiometrically reduced by limiting the concentrations of Mg2+. Our results suggest that a complex of citrate and Mg2+ is the transported substrate. The coupled transport of citrate and Mg2+ does function normally in a Co2+-resistant mutant of B. subtilis which is defective in another transport system of high affinity for Mg2+. The efflux of phosphate from B. subtilis cells during uptake of citrate is too slow to account for electroneutrality of the system. No cotransport of K+ and citrate can be demonstrated. Valinomycin inhibits the citrate-Mg2+ transport activity, which can be restored by the addition of external K+. In a potassium-retention mutant of B. subtilis, citrate transport activity decreases in parallel with the loss of K+ from these cells. Thus a high concentration of K+ inside the cells is necessary for the proper function of the citrate-Mg2+ transport system.