Figure 3: Transport kinetics of Soa1 and ASTA.

Cells were starved for sulfur, buffered in Bis-Tris pH 6.5 and transport rates were measured by short-term exposure to different concentrations of radiolabelled isethionate (a), taurine (b) and sulfate (c,d). For low-affinity sulfate transport, the concentration was kept constant at 40 mM and transport was measured for 1 and 10 min (c). The affinity constants calculated using the reciprocal approximation of the Hanes–Woolf plots shown as insets (K_m of Soa1: 103.8 μM and K_m of astA: 8.5 μM) are slightly different from the more precise nonlinear fit to the Michalis–Menten equation already reported (K_m of Soa1: 116.9 μM and K_m of astA: 7.5 μM). The pH profiles were assayed in different buffers with adjusted pH, while keeping the amount of isethionate (500 μM) and sulfate (200 μM) constant (A,D). H⁺-symport activity was tested by exposing cells to the proton gradient decoupling agent CCCP for 10 min before transport measurements (a,d). All experiments were carried out in triplicate from aliquots of the same cell preparation assayed independently with the mean shown and error bars indicating s.d. In many cases, the error is so small that the error bars are not visible.