A single amino acid substitution (Glu134‐‐>Ala) in NhaR1 increases the inducibility by Na+ of the product of nhaA, a Na+/H+ antiporter gene in Escherichia coli

The mutation nhaAup (antup) has now been identified as a Glu134 to Ala substitution in NhaR and designated nhaR1. This was demonstrated by sequence analysis showing that the mutant contains a wild‐type nhaA but nhaR1 instead of nhaR and by the finding that nhaR1 cloned in a plasmid confers the NhaAup phenotype. Na+ (107 mM) increases by 5‐ to 10‐fold the level of nhaA transcripts, similar to the effect on the NhaR‐mediated expression of a nhaA‘‐’lacZ fusion. These results are in agreement with the notion that nhaR is a positive regulator which controls Na(+)‐dependent transcription of nhaA. The promoter region of nhaR and nhaR1 was found to reside within the BglII‐BamHI fragment of the C‐terminal sequences of nhaA. The mutation nhaR1, while increasing dramatically the level of transcription, reduces the requirement for Na+ by 3‐ to 5‐fold both for nhaA transcription and for the nhaR1‐mediated expression of nhaA‘‐’lacZ fusion. NhaR1, like NhaR, binds specifically to the promoter region of nhaA. However, at equal protein concentration NhaR1 binds more DNA and the NhaR1‐DNA complex shows higher mobility than that of NhaR‐DNA, suggesting the existence of two different binding complexes. Yet in this assay the DNA binding pattern of neither NhaR nor NhaR1 was affected by the addition of Na+. The possible relevance of these two DNA‐binding complexes to the Na(+)‐induced NhaR‐mediated expression is discussed.