Fig. 5.

EmrE-W63G but not EmrE is capable of membrane potential driven [³H]putrescine uptake when reconstituted into proteoliposomes. Proteoliposomes reconstituted with purified EmrE or EmrE-W63G loaded with 150 mM KCl and 20 mM K–Hepes (pH 7.5) were diluted into a medium containing 150 mM NaCl, 20 mM Na–Hepes (pH 7.5), 100 μM [³H]putrescine, and 10 μM of the potassium ionophore valinomycin (to generate the membrane potential). Transport was then measured over time as described in Materials and Methods. (A) Squares: EmrE-W63G; circles: EmrE B. The same transport reaction was carried out with EmrE-W63G proteoliposomes for 2 min as mentioned above but also including increasing concentrations of agmatine (circles), BTP (squares), or erythromycin (triangles).