A stable Na+/H+ antiporter of thermophilic bacterium PS3

As a first step in the isolation of a stable Na+/H+ antiporter, its reaction in sonicated membrane vesicles of thermophilic bacterium PS3 has been characterized. The sonicated vesicles showed quenching of quinacrine fluorescence in either NADH oxidation or ATP hydrolysis. The quenching was reversed by the addition of Na+, Li+, Mn2+, Cd2+, and Co2+, but not of choline+ or Ca2+, regardless of their counter anions. 22Na+ was taken up into the vesicles by NADH oxidation, and the 22Na+ uptake was inhibited by the addition of an uncoupler. H+ release was observed on addition of Na+ to sonicated vesicles. The magnitude of the pH difference across the membrane induced by NADH oxidation was constant at pH 7.0 to 9.1, but the Na+/H+ antiport was affected by the pH of the medium (optimum pH-8.5). The Km's of the antiporter for Na+ and Li+ were 2.5 and 0.1 mM, respectively, but the Vmax values for the two ions were the same at pH 8.0. In the presence of Li+, no further decrease of fluorescence quenching was observed on addition of Na+ and vice versa. The Na+/H+ antiporter activity in PS3 was stable at 70 degrees C, and the optimum temperature for activity was 55-60 degrees C. In contrast to mesophilic cation/H+ antiporters, this antiporter was not inhibited by a thiol reagent.