Known and predicted binding ion sites in the ion transporter superfamily fold. Two Na⁺-binding sites have been reported based on the experimental electron density; these ions are shown as opaque purple spheres, and residues coordinating those ions directly are indicated by labels in bold font. (A) In VcINDY (PDB accession no. 4F35), a density was identified at the region labeled Na2. (B) In YdaH (PDB accession no. 4R0C), a density was identified at the region labeled Na3. The corresponding sites in VcINDY (Na3) and YdaH (Na2), shown as transparent spheres, were predicted after superposing YdaH on VcINDY with Fr-TM-align. Putative coordinating residues (Asn, Gln, Asp, Glu, Thr, or Ser within 8 Å of the ion) for the predicted YdaH-Na2 and VcINDY-Na3 site regions are indicated with spheres and labeled. The ion nomenclature follows that adopted previously for NaPi-II (see Results). The protein is shown as ribbons, and residues of interest are highlighted using spheres at the position of the Cα atom. (C) The equivalent region in MtrF (PDB accession no. 4R1I), which is a putative H⁺-coupled transporter. (D) Structural model of human NaPi-IIa, with ions placed according to the results from VcINDY and YdaH shown as purple spheres. For NaPi-IIa, biochemical and electrophysiological evidence supports a role in phosphate or sodium binding for the residues shown as spheres (*). During the transport cycle of NaPi-IIa, an additional sodium binds before Na2 and Na3, at a site named Na1 (not depicted). The structures of VcINDY, YdaH, and MtrF are oriented with the extracellular side toward the top of the page, whereas NaPi-IIa is oriented with the cytoplasmic side toward the top of the page, because it inserts in the membrane in the opposite direction from the other transporters.