Extended Data Figure 7: The extracellular pathway.

a, The extracellular fraction of the E2–AlF₄⁻ crystal structure. Functionally important residues are shown as sticks, and the protein is coloured as in Fig. 1a. The final 2F_o − F_c electron density is contoured at 1σ. The view is equivalent to the one in Fig. 3d. b, Dynamics of E202 in a 60-ns molecular dynamics simulation of the E2–BeF₃⁻ structure in a dioleoylphosphatidylcholine (DOPC) membrane in the absence of zinc. Selected residues are shown as sticks. Representative E202 conformations were captured at 16, 25 and 30 ns from snapshots aligned according to backbone Cαs of M1–M4. The orientation of E202 at 16 ns resembles how this side chain appears in the E2–AlF₄⁻ state, while the flexibility observed throughout the simulation agrees with the observed poor electron density of the side chain in the E2–BeF₃⁻ state (see Fig. 3b). Note that there are two distorted lipids at the release pathway that may assist in Zn²⁺ release (vdW spheres represent lipid phosphates). c, Distance between the centre of mass of the Cδ of the E202 side chain and the N_Z of the K693 side chain during the 60-ns simulations of the E2–AlF₄⁻ and E2–BeF₃⁻ S. sonnei ZntA structures in the absence of zinc, as a running average over five consecutive frames of each trajectory. d, The release pathway and accompanying protein interactions experienced by Zn²⁺ in a steered molecular dynamics simulation originating from the centre of mass of residues C392, C394 and D714. The transmembrane domain, lipid phosphates and water within 7 Å of the protein are coloured as in b. e, The number of Zn²⁺–protein interactions with a 5 Å cut-off during steered molecular dynamics (SMD) simulations. Error bars correspond to counts from ten independent simulations with pulling speeds on Zn²⁺ of 10–20 Å ns⁻¹.