Characterization of sulfate transport in Desulfovibrio desulfuricans

Abstract

Uptake of ³⁵S-labelled sulfate was studied with a new isolate of Desulfovibrio desulfuricans, strain CSN. Micromolar additions of sulfate (1–10 μM or nmol/mg protein) to cell suspensions incubated in 150 mM KCl at-1°C were almost completely taken up and accumulated about 5,000-fold. Accumulation was not influenced by incubation in NaCl instead of KCl, by acidic pH (5.5) or by incubation under air for 10 min. In alkaline milieu (pH 8.5), after prolonged contact with air (2 h), or after growth with excess sulfate or thiosulfate as electron acceptor, the amount taken up was diminished approximately by half. Pasteurization inhibited sulfate uptake completely. With increasing concentrations of added sulfate (0.1 to 2.5 mM) the intracellular concentration increased only slowly up to 25 mM, and the accumulation factor decreased down to 8. Sulfate transport was reversible. Accumulated sulfate was rapidly lost from the cells after addition of excess non-labelled sulfate or after addition of the uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP). The ATPase inhibitor dicyclohexylcarbodiimide (DCCD) specifically inhibited sulfate reduction but had no immediate influence on sulfate accumulation. Addition of the phosphate analogue arsenate (5 mM) was without effect. These results were not in favour of an ATP-dependent transport system. The K⁺-H⁺-antiporter nigericin (in 150 mM KCl) and the Na⁺-H⁺-antiporter monensin (in 150 mM NaCl) caused partial inhibition of sulfate accumulation, whereas the K⁺-transporter valinomycin (in 150 mM KCl) and the Na⁺-H⁺ exchange inhibitor amiloride (2 mM) were without effect. The permeant thiocyanate anion (150 mM) inhibited sulfate uptake by 60% at pH 7, and completely at pH 8.5. Although the effects of the different ionophores on the chemiosmotic gradients have not been studied so far, the results indicated that probably both, ΔpH and ΔΨ drive sulfate accumulation and that sulfate is taken up electrogenically in symport with more than 2 protons. The structural sulfate analogues tungstate and molybdate (0.1 mM, each) did not affect sulfate accumulation, although molybdate inhibited sulfate reduction. Chromate completely blocked both of these activities. Sulfite and selenite caused little or no decrease of sulfate accumulation, whereas with thiosulfate and selenate significant inhibition was observed.