Structures. (A, left) Torpedo AChR (PDB accession code 2BG9): α subunit, tan; γ subunit, white. The TBS is ∼50 Å from the gate. (right) AChR α subunit diliganded gating phi values mapped onto Caenorhabditis elegans GluCl (PDB accession code 3RIF); colors assigned by statistical criteria (Fig. 6 A; Purohit et al., 2013). Amino acids at the TBS and in αM2M3 have phi ∼1 (purple) and are separated by a domain of phi ∼0.8 residues. For clarity, some amino acids were removed at the ECD–TMD interface and M3. (B, top) Ligand-binding site of the Lymnaea stagnalis ACh-binding protein (PDB accession code 3WIP; AChR numbers). In adult AChRs, affinity is mainly determined by αY190, αY198, and αW149 (green), but in fetal AChRs, αY93 and γW55 also contribute (yellow). (bottom) ECD–TMD interface of human α4β2 AChRs (PDB accession code 5KXI). (inset) In GluCl, the M2 helix is displaced upward relative to M1 in O versus C (PDB accession codes 3RIF and 4TNV). (C) M2M3 sequence alignments. *, αP265 in mouse muscle AChRs. Highlighted residues are loop.