FIGURE 6.

Activation of ENaC by subtilisin cleavage of α. A, subtilisin caused marked time-dependent activation of the amiloride-sensitive conductance in α/β/γ-expressing oocytes. Stimulation was dose-dependent with activation observed at levels as low as 2 ng/ml (B). Activation peaked at 15 min at 50 ng/ml subtilisin. In this group of experiments 50 ng/ml subtilisin activated αWT by ∼6-fold and αFM by ∼35-fold. C, both activated channels reached similar absolute values, indicating the same mechanism of activation (p < 0.01 between the conductance of α/β/γ and αFM/β/γ before subtilisin and p > 0.05 after subtilisin). This effect is consistent with the biochemical data summarized in Fig. 4 and the in vitro cleavage data shown below. Subtilisin did not affect oocyte endogenous currents (control oocytes are not shown, but note the absence of changes of amiloride-sensitive currents before and after subtilisin). n = 5 for B and n = 10 for C.