Ligand pathways and binding sites in the three inward-facing structures. Cartoon representation of the MtrF (A), YdaH (B), and VcINDY (C) dimer structures viewed along the membrane plane. The subunits on the right are colored with separate colors for each helix. Ligands are shown as spheres, with their presumed pathways illustrated using arrows. The position of the dimers in the membrane was determined for the x-ray structures using the Orientations of Proteins in Membranes (OPM) server. Each of the three transporters forms an upside-down bowl-shaped structure with a concave aqueous basin facing the intracellular side. VcINDY has a citrate molecule and a Na⁺ ion modeled in the binding site of each protomer, whereas YdaH has a Na⁺ ion in each protomer. No substrate was detected in the MtrF structure. To the right of each dimer, the transport domain is shown with a surface representation of the binding pocket in white, indicating that the binding sites vary in size in the presence of substrates. (D) Comparison of the tilt angle of TM5b in VcINDY (light blue and pink) with either TM3b of YdaH (left; dark blue and red) or TM3b of MtrF (right; dark blue and red). The binding pockets were superimposed using the first three helix turns of HP2b, and the angle of TM3b relative to TM5b was calculated. The position of the ligands is shown for VcINDY (transparent spheres) and YdaH (purple sphere).