A Phe at position D455 stabilizes the lower pore and rescues destabilizing nonfunctional lower pore mutants. (A) Summary of normalized current, τd, and recovery ratio of various D455 substitutions. (B) Representative examples of current traces of D455 substitutions. (C) Summary of normalized current, τd, and recovery ratio of double mutants (D455F and an additional substitution in the lower pore). (D) Representative examples of current traces of the double mutants. Data are presented as mean ± SD of three independent experiments for at least five Xenopus oocytes (n = 5–10) for each construct tested. All oocytes were incubated at least 30 s (30–40 s) in preconditioning buffer (pH 7.4) for channel recovery before the next activation. Recovery ratio (expressed in percent) was calculated as Iacti(n + 1)/Iacti(n). Individual measurements are shown in Data S2.