FIGURE 10.

Effect of DEPC on the proton transport by hASCT2 reconstituted in proteoliposomes. Purified hASCT2 was reconstituted in proteoliposomes buffered at pH 7.0, containing 10 mM glutamine and 0.1 mM pyranine, as described in materials and methods. After reconstitution, proteoliposomes were incubated with 10 mM DEPC (gray line) or ethanol (black line) for 5 min at 23°C and, then were subjected to sephadex-G75 chromatography. The fluorescence measurement was started by adding 150 μL proteoliposomes to transport buffer (up to 3 mL final volume) composed by 100 mM Na-gluconate and Hepes Tris 20 mM at pH 7.0. After ∼2 min as indicated by the arrow, 5 mM glutamate was added to the sample and fluorescence was recorded. The fluorescence signal is indicated as arbitrary units (AU). Results are representative of three independent experiments.