MOLECULAR PHYSIOLOGY OF THE NA+ H+ ANTIPORTER IN ESCHERICHIA-COLI/

All living cells maintain an inwardly directed Na+ gradient and a cons tant intracellular pH. Na+/H+ antiporters have been assigned an essent ial role in these homeostatic mechanisms in all cells. In Escherichia coli, two Na+/H+ antiporter genes, nhaA and nhaB, have been cloned. De letion of either one or both showed that NhaA is essential for adaptat ion to high salinity, for growth at alkaline pH in the presence of Nat and for challenging Li+ toxicity. NhaB confers tolerance to low level s of Na+ and becomes essential when the activity of NhaA limits growth . The adaptive response to Na+ is mediated by the positive regulator n haR, which transduces the signal (intracellular Na+) to expression of the nhaA gene. We have identified Glu-134 of NhaR as part of the 'Nasensor' of NhaA. In agreement with the role of NhaA in pH homeostasis, its Na+-dependent expression is enhanced at alkaline pH. Reconstituti on of pure NhaA and NhaB in proteoliposomes demonstrates that, whereas both are electrogenic (the H+/Na+ stoichiometry of NhaA is 2), only N haA is pH-dependent, increasing its activity 1000-fold between pH7 and 8.5. Mutating all the histidines of NhaA shows that His-226 is part o f the 'pH sensor' of NhaA.