Differential effects of mutations on the transport properties of the Na

Background: NhaA of Escherichia coli can serve as an example for pH regulation and transport mechanism in Na + /H + -exchangers. Results: Different mutations alter Na + and/or H + binding or conformational dynamics. Conclusions: Mutations in different segments of the protein change its pH profile by affecting different aspects of its transport mechanism. Significance: Single site mutations and kinetic analysis reveal determinants of Na + /H + exchanger mechanism. SUMMARY Na + /H + -antiporters show a marked pH dependence which is important for their physiological function in eukaryotic and prokaryotic cells. In NhaA, the Escherichia coli Na + /H + -antiporter, specific single site mutations modulating the pH profile of the transporter have been described in the past. To clarify the mechanism by which these mutations influence the pH dependence of NhaA the substrate dependence of the kinetics of selected NhaA variants was electrophysiologically investigated and analyzed with a kinetic model. It is shown that the mutations affect NhaA activity in quite different ways by changing (i) the properties of the binding site or (ii) the dynamics of the transporter. In the first case