FIGURE 3.
Effect on conductance block of LFN of adding a single charged residue into the Phe clamp. A, macroscopic conductance was measured at symmetrical pH of 5.5 and Δψ = +5 mV for [WT]7 (black circles), [WT]6[F427K]1 (dark blue circles), [WT]6[F427R]1 (light blue circles), [WT]6[F427H]1 (green circles), [WT]6[F427D]1 (orange circles), and [WT]6[F427E]1 (purple circles). The channels were blocked with 10 nm LFN, and then dissociation of LFN was monitored over time during perfusion of the cis compartment. B, representative single-channel currents of heteroheptameric PA wild type and mutant channel activity blocked by the addition of 1 μg/ml LFN to the cis compartment, recorded at Δψ = +20 mV in 0.11 m KCl. C and O denote the closed and open states, respectively. C, translocation of LFN through macroscopic PA channels. At time 0, translocation was initiated by adding 2 m KOH to the trans compartment to raise the pH to 7.2; there was an ∼20-s mixing delay in this system with both compartments continuously stirred. Representative data are shown from n ≥ 3 trials.