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Abstract

We examined substrate-induced conformational changes in MjNhaP1, an archaeal electroneutral Na⁺/H⁺-antiporter resembling the human antiporter NHE1, by electron crystallography of 2D crystals in a range of physiological pH and Na⁺ conditions. In the absence of sodium, changes in pH had no major effect. By contrast, changes in Nav concentration caused a marked conformational change that was largely pH-independent. Crystallographically determined, apparent dissociation constants indicated ∼10-fold stronger Na⁺ binding at pH 8 than at pH 4, consistent with substrate competition for a common ion-binding site. Projection difference maps indicated helix movements by about 2 Å in the 6-helix bundle region of MjNhaP1 that is thought to contain the ion translocation site. We propose that these movements convert the antiporter from the proton-bound, outward-open state to the Na⁺-bound, inward-open state. Oscillation between the two states would result in rapid Na⁺/H⁺ antiport