Gütter C , Benndorf K , Zimmer T .

	Figure 1. Schematic representation of hNav1.5 and the N-terminal mutations investigated in this study. (A) Proposed hNav1.5 topology. Affected residues are indicated in white (LQT3) and light grey (BrS). The schematic structure also highlights some important structural features (DI to DIV—domain I to IV, IFM—residues isoleucine, phenylalanine, and methionine of the inactivation gate, IQ—calmodulin binding motif, EF—Ca++ binding EF hand domain). (B) Alignment of the N-terminal sequences of human Nav1.1—Nav1.9. Mutations associated with LQT3 or BrS are underlined or indicated in italics, respectively. Most of the eight affected residues are conserved among the Nav1 subfamily. Residues at position 35 are variable, and position 125 is occupied by isoleucine in all other human Na+ channels. All eight residues affected by a SCN5A mutation are identical in Nav1.5 of human, rat, mouse, and dog (not shown). The N-terminal region of the first putative membrane spanning segment DI-S1 is indicated in light grey. References: G9V (Millat et al., 2006), R18W (Tester et al., 2005), R18Q (Kapplinger et al., 2010), R27H (Priori et al., 2002), G35S (Levy-Nissenbaum et al., 2001), V95I (Liang et al., 2006), R104Q (Levy-Nissenbaum et al., 2001), V125L (Tester et al., 2005), K126E (Vatta et al., 2002a).
	Figure 2. Whole-cell Na+ currents upon expression in HEK293 of hNav1.5 mutant channels associated with LQT3. (A) Current families. Currents were elicited by test potentials from −80 mV to various test pulses in 5 or 10 mV increments at a pulsing frequency of 1.0 Hz. (B) Persistent currents at −20 mV. The non-inactivating current fraction was similarly small in both wild-type and mutant hNav1.5 channels. For individual values see Table 1. Mutant ΔKQP channels were used as a positive control.
	Figure 3. Electrophysiological properties in HEK293 cells of mutant hNav1.5 channels associated with LQT3. (A) Inactivation time constants τh (ms) as function of voltage. At −50 mV both R18W and V125L channels inactivated more slowly compared to hNav1.5 (*indicates p < 0.05). G9V channel inactivation was indistinguishable from hNav1.5 (data not shown). (B) Steady-state activation and inactivation in V125L channels. The figure shows the mean of 4 representative measurements for each. (C) Window current in hNav1.5 (grey area, solid lines) and V125L (dark grey area, dotted lines). (D) Recovery from inactivation was accelerated in V125L (* indicates p < 0.05 vs. hNav1.5). For individual values see Table 2.
	Figure 4. Whole-cell Na+ currents upon expression in HEK293 of hNav1.5 mutant channels associated with BrS. Currents were elicited from −80 mV to various test pulses in 5 or 10 mV increments at a pulsing frequency of 1.0 Hz (holding potential −120 mV). For average peak current densities see Table 1. Expression of R104Q did not result in functional channels in the mammalian expression system.
	Figure 5. Electrophysiological properties in HEK293 cells of mutant hNav1.5 channels associated with BrS. (A) Inactivation time constants τh (ms) as function of voltage. K126E channels inactivated more slowly at −50 mV (**), R27H inactivated more slowly from −50 to −25 mV (*), when compared to hNav1.5. R18Q, G35S, and V95I channels were indistinguishable from hNav1.5 (data not shown). (B) Steady-state activation and steady-state inactivation curves. Mid-activation potentials (Vm) were significantly shifted towards depolarized potentials in both R27H and K126E. Mid-inactivation potential Vh was shifted only in K126E. The figure was drawn using the mean of 4 representative measurements for each. Vm and Vh of the other mutant channels, R18Q, G35S and V95I, were indistinguishable from the respective hNav1.5 values. For data and statistics see Table 2.
	Figure 6. Electrophysiological properties of R104Q in Xenopus oocytes. (A) Whole-cell Na+ currents. Peak current amplitudes were significantly reduced in R104Q (see Table 1). Currents were elicited from −80 mV to various test pulses in 5 or 10 mV increments at a pulsing frequency of 1.0 Hz (holding potential −120 mV). (B) Steady-state activation and inactivation curves. In R104Q, steady-state availability was significantly reduced. (C) Recovery from inactivation was slower in R104Q (* indicates p < 0.05 vs. hNav1.5). For individual values see Table 3.

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