Figure 2.

d-[³H]Aspartate transport in proteoliposomes containing wild-type GLT-1 or transporters with mutations at positions 440 and/or 443. HeLa cells were infected with vaccinia/T7 recombinant virus and transfected with wild-type or mutant transporters. Subsequently, the transporters were solubilized and reconstituted as described in Materials and Methods. The internal medium contained 0.12 M potassium phosphate, pH 7.4. Net influx of d-[³H]aspartate was measured at the indicated times using 0.15 M of either NaCl (■), LiCl (●), or choline chloride (not shown, but indistinguishable from the abscissa) and 2.5 μM of valinomycin. (A) Wild type (WT) and S440G–S443Q. (B) S440G and S443Q. (C) S440G–S443G and S443G. For each time point the data are expressed as percentage of the value in sodium-containing medium after 10 min. These values were 2,448, 833, 803, 1,508, 450, and 2,302 pmol/mg protein for wild type, S440G–S443Q, S440G, S443Q, S440G–S443G, and S443G transporters, respectively.