A gene encoding an Na+/H+ antiporter was cloned from chromosomal DNA of Vibrio parahaemolyticus, a slightly halophilic bacterium, and expressed in Escherichia coli cells. The gene enabled mutant E. coli cells, which were unable to grow in the presence of 10 mM LiCl (or 0.2 M NaCl) because of the lack of major Na+(Li+)/H+ antiporters, to grow under such conditions. We detected Na+/H+ antiport activity due to the gene in membrane vesicles prepared from E. coli cells that harbored the plasmid carrying the gene. Li+ was also a substrate for this antiporter. Activity of this antiporter was pH-dependent with highest activity at pH 8.5 to 9 and no activity at 7.0 to 7.5. Restriction mapping and a Southern blot analysis revealed that the cloned gene was different from the nhaA and the nhaB of V. parahaemolyticus. We designated the gene nhaD. The gene was sequenced, and the amino acid sequence of the NhaD protein was deduced. The NhaD is a unique Na+/H+ antiporter with respect to the primary structure compared with known Na+/H+ antiporters.
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