Effect of α-R205A/R231A, β-, and γ-ENaC mutant on I_Na and fragments stimulated by CAP2. (A and B) Surface biotinylated α-ENaC N-terminal and C-terminal fragments were visualized by Western blot analysis using anti-HA or anti-V5 monoclonal antibodies. (A) Many nonspecific (*, NS) bands evident in film overexposed to reveal the fate of the furin (32 kD) and novel fragments (19 and 17 kD). FL, full-length. Lane 1, uninjected eggs; lane 2, WT ENaC alone; lane 3, WT ENaC plus CAP2; lane 4, α-R205A/R231A, β-, and γ-ENaC alone; lane 5, α-R205A/R231A, β-, and γ-ENaC plus CAP2. (C) CAP2-mediated I_Na of WT or mutant channels were measured as described above. Batches of oocytes were extracted from five different frogs (n = 31). Results are expressed as the means ± SE. * and **, P < 0.0001. Statistical significance was determined using an unpaired Student's t test.