Figure 6: Residues outside of the DII paddle motif contribute to rDc1a binding.
(a) Sequence alignment of the DII S1–S2 regions of BgNav1 and PaNav1 (See Supplementary Fig. 4 for more details). In the extracellular loop connecting S1 to S2, all residues are identical except the H805Y and D812E substitutions (grey background). The T797M substitution within S1 is unlikely to be toxin-accessible. (b) Gating properties of the mutant BgNav1YE channel. Shown are the normalized deduced conductance (G)–voltage (filled circles) (G/Gmax) and steady-state inactivation (open circles; I/Imax) relationships. Descriptive values can be found in the Results section. Currents were elicited by 5 mV step depolarizations from a holding voltage of −90 mV or −100 mV, respectively. n=3–5, and error bars represent s.e.m. (c) Affinity measure for rDc1a interaction with BgNav1YE (red) compared with WT BgNav1 (grey). Concentration dependence for toxin-induced current potentiation (as determined by shifts in V1/2) is shown. It is clear that alterations in channel opening on rDc1a exposure are less pronounced with the BgNav1YE mutant when compared with WT BgNav1. n=5 and error bars represent s.e.m. Inset shows premature channel opening after addition of 100 nM rDc1a to WT BgNav1 (grey) whereas BgNav1YE is not yet affected (red). Current traces were evoked by a 50-ms depolarization to −55 mV from a holding potential of −90 mV.
