Fig. 3: Key residues involved in the conformational change of hOCT1.

a Overall structures of outward facing (hOCT1-S1) (orange) and inward facing (hOCT1-S2) (cyan) hOCT1 with a close-up view of the YER motif. b Top view of hOCT1 in the outward facing (hOCT1-S1) (orange) and inward facing (hOCT1-S2) (cyan) conformations reveals that the YER motif links change in the central pocket during the conformational rearrangement of the CTD. Black arrows indicate YER motif side chain rearrangements. Red arrows indicate movement of TM7 and TM10 that directly interact with the YER motif. Blue arrows indicate the movement of TM9, TM11 and TM12 that are indirectly connected to YER motif. c Sequence alignment suggests that the YER motif (highlighted in red) is conserved in SLC22A1-8. d Mutating any YER motif residue impaired metformin (green) and MPP⁺ (orange) uptake activities of hOCT1. Data are normalized to wild-type hOCT1 and are shown as mean ± SEM of four independent experiments. e W354 interacts with N453 in the inward occluded (hOCT1-S2) (cyan) state, while the interaction is broken by the inward open (hOCT1-apo1) (magenta) state. f Bottom view of hOCT1 in the inward occluded (hOCT1-S2) (cyan) and inward open (hOCT1-apo1) (magenta) states. The side chain rotation of W354 and the movement of N453 are associated with the inward channel opening. Black arrows indicate the movement of W354 and N453. Red arrow indicates the displacement of TM10 that directly links with N453. Blue arrows indicate the outward displacement of TM8 and TM11 that is influenced by TM10. g The W354A mutation impairs metformin (green) and MPP⁺ (orange) uptake by hOCT1, while the N453A mutation only impairs the metformin (green), but not the MPP⁺ (orange), uptake activity of hOCT1. Data are normalized to wild-type hOCT1 and are shown as mean ± SEM of four independent experiments.