FIGURE 3.

Na⁺(Li⁺, K⁺)/H⁺ antiport activity by MdrP and its activity pH profile. Na⁺(Li⁺, K⁺)/H⁺ antiport activity was determined in everted membrane vesicles prepared from cells of E. coli KNabc/pET-mdrP (to the left) or KNabc/pET19b (to the right) by the French pressure cell method. The optimal antiport activity at pH 9.0 for Na⁺/H⁺ (A) and Li⁺/H⁺ (B), and at pH 8.5 for K⁺/H⁺ (C) were shown as the representatives of each of them. At the time points indicated by downward arrows, Tris-D-lactic acid (final concentration at 5 mM) was added to the assay mixture to initiate fluorescence quenching. At the time points indicated by upward arrows, NaCl (final concentration at 5 mM), LiCl (final concentration at 5 mM) or Na-free KCl (final concentration at 5 mM) was added to the assay mixture, respectively. Fluorescence quenching is shown in arbitrary units (AU). The antiport activity pH profile (D) for Na⁺/H⁺ (filled circle), Li⁺/H⁺ (filled square) and K⁺/H⁺ (filled triangle) were also plotted at the indicated pH values. The wavelength of excitation light was 492 nm and fluorescence was monitored at 526 nm. Each value point represents the average of three independent determinations.