Fig. 1: Conformational states of ECF transporters.

Cryo-EM maps of the nucleotide-bound ECF-FolT2_ATP complex (a), the nucleotide-free wild-type solitary ECF module (b), and the ATP-bound mutant solitary ECF module 2EQ_cryo (c), all in lipid nanodiscs, viewed from the membrane plane onto the docking site of FolT2 (top) and rotated by 90° onto the site of the cytoplasmic substrate exit pathway (grey arrow) for folate from the cavity in FolT2. The shape of the coupling helices (CH2 and CH3) is indicated by white dotted lines with the terminal triangles indicating the conserved X-Arg-X anchor motifs. Bound nucleotides are indicated by white contours (solid lines - located towards the front, dashed lines - towards the back) and labels. The green arrows in (a) and (b) indicate the direction in which the structural elements have moved relative to (c). Similarly, the purple arrows indicate the opposite movement. Cross sections of the ECF-FolT2_ATP complex (d), the wild-type solitary ECF module (e), and the mutant solitary ECF module 2EQ_cryo (f) revealing the shape of the interaction surface of the coupling helices and the S-component. Models are shown in surface representation. The schematic representations in the boxes indicate the position of the slice through and the zoomed in region of the cross section. An outline of FolT2 is shown highlighting a good fit without clashes with the ECF module in the ATPase open conformation (e) or clashes with the ECF module in the ATPase closed conformation (f). f The surfaces of the clashes are shaded in yellow and are indicated with a lightning symbol. a–f The approximate boundaries (grey lines) of the membrane and the extracellular (EC) and intracellular (IC) sides are indicated. The individual subunits/regions are coloured as follows: EcfA in red, EcfA’ in orange, EcfT coupling helices (CHs) in dark blue, EcfT transmembrane domain (TMs) in light blue, and the S-component FolT2 in yellow.