Fig. 5. Conductances, lifetimes and overall membrane activities of RtxA and proRtxA on planar lipid membranes.

a Representative single-pore recordings of asolectin/decane:butanol (9:1) membranes in the presence of 1 nM purified recombinant RtxA and proRtxA. The aqueous phase contained 150 mM KCl, 10 mM Tris-HCl (pH 7.4), 2 mM CaCl₂; the applied voltage was 50 mV; the temperature was 25 °C and the recording was filtered at 10 Hz. The numbers above the recordings represent lifetimes. b Kernel density estimation (KDE) of single-pore conductances of RtxA and proRtxA (>500 events) were calculated from single-pore recordings acquired on several different asolectin membranes under the same conditions as in (a). The results are summarized in Table 1, and the numbers in each KDE represent the most frequent values of three well-distinguishable single-pore conductance states. c For lifetime determination, approximately 400 individual pore openings were recorded on several different asolectin membranes with 0.25, 0.5, and 1 nM RtxA or proRtxA (all other parameters were the same as in (a)), and the KDE of dwell times was fitted with a double-exponential function. The error estimates of lifetimes were obtained by bootstrap analysis. The results are summarized in Table 1, and the numbers in each graph represent the most frequent values of two well-distinguishable lifetimes. d Overall membrane activity of the purified RtxA and proRtxA proteins on asolectin/decane:butanol (9:1) membranes using the same conditions as in (a)