A single residue differentiates between human cardiac and skeletal muscle Na+ channel slow inactivation.
Slow inactivation determines the availability of voltage-gated sodium channels during prolonged depolarization. Slow inactivation in hNa(V)1.4 channels occurs with a higher probability than hNa(V)1.5 sodium channels; however, the precise molecular mechanism for this difference remains unclear. Using the macropatch technique we show that the DII S5-S6 p-region uniquely confers the probability of slow inactivation from parental hNa(V)1.5 and hNa(V)1.4 channels into chimerical constructs expressed in Xenopus oocytes. Site-directed mutagenesis was used to test whether a specific region within DII S5-S6 controls the probability of slow inactivation. We found that substituting V754 in hNa(V)1.4 with isoleucine from the corresponding position (891) in hNa(V)1.5 produced steady-state slow inactivation statistically indistinguishable from that in wild-type hNa(V)1.5 channels, whereas other mutations have little or no effect on slow inactivation. This result indicates that residues V754 in hNa(V)1.4 and I891in hNa(V)1.5 are unique in determining the probability of slow inactivation characteristic of these isoforms. Exchanging S5-S6 linkers between hNa(V)1.4 and hNa(V)1.5 channels had no consistent effect on the voltage-dependent slow time inactivation constants [tau(V)]. This suggests that the molecular structures regulating rates of entry into and exit from the slow inactivated state are different from those controlling the steady-state probability and reside outside the p-regions.
Vilin
Y Y
YY
Department of Biology, Utah State University, Logan, Utah 84322, USA.
Fujimoto
E
E
Ruben
P C
PC
D000595:Amino Acid Sequence; D000818:Animals; D004594:Electrophysiology; D006801:Humans; D007532:Isoleucine / Q000737:chemistry; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009206:Myocardium / Q000378:metabolism*; D009865:Oocytes / Q000378:metabolism; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D015222:Sodium Channels / Q000737:chemistry* / Q000235:genetics; D014633:Valine / Q000737:chemistry; D014981:Xenopus
D020033:Protein Isoforms; D015222:Sodium Channels; D007532:Isoleucine; D014633:Valine