Essential N- and C-terminal residues for hASIC1a function. (A) Normalized peak currents of indicated hASIC1a N-terminal truncations measured from Xenopus oocytes injected with cRNA of WT hASIC1a or with truncations examined by TEVC. Normalized peak currents are presented as mean ± SD of three independent experiments of at least 10 oocytes (n = 10–25) for each construct. One-way ANOVA post hoc Dunnett’s F test 49.2; ***, P < 0.0001. Individual measurements are shown in Data S2 for A and B. (B) Normalized peak currents of indicated hASIC1a C-terminal truncations measured and analyzed as in A. ***, P < 0.0001; *, P = 0.01. (C) Sequence alignment of amino acids encoding the lower pore of ASICs. From various species using Lasergene DNASTAR-MegAlign software. Highlighted residues were examined in this work. c, chicken; h, human; l, lamprey; r, rat; s, shark; x, Xenopus; z, zebrafish. (D–F) Pore opening repositions functionally essential residues in the lower pore of hASIC1a. (D) Side view of the predicted hASIC1a in the open conformation (blue) superimposed onto the predicted hASIC1a in the resting conformation (tan). (E) Enlarged view of the TM domains and the reentrant loop. (F) View of the TM domains from the intracellular side shows rotation of TM1 and TM2 around the threefold axis and the predicted repositioning of the reentrant loop from the closed (tan) to the open (blue) conformation.