Intrinsic voltage dependence of the epithelial Na+ channel is masked by a conserved transmembrane domain tryptophan. J Biol Chem 2009 Sep 18;284(38):25512-21
Date
07/22/2009Pubmed ID
19620245Pubmed Central ID
PMC2757952DOI
10.1074/jbc.M109.015917Scopus ID
2-s2.0-70350023553 (requires institutional sign-in at Scopus site) 11 CitationsAbstract
Tryptophan residues critical to function are frequently located at the lipid-water interface of transmembrane domains. All members of the epithelial Na+ channel (ENaC)/Degenerin (Deg) channel superfamily contain an absolutely conserved Trp at the base of their first transmembrane domain. Here, we test the importance of this conserved Trp to ENaC/Deg function. Targeted substitution of this Trp in mouse ENaC and rat ASIC subunits decrease channel activity. Differential substitution with distinct amino acids in alpha-mENaC shows that it is loss of this critical Trp rather than introduction of residues having novel properties that changes channel activity. Surprisingly, Trp substitution unmasks voltage sensitivity. Mutant ENaC has increased steady-state activity at hyperpolarizing compared with depolarizing potentials associated with transient activation and deactivation times, respectively. The times of activation and deactivation change 1 ms/mV in a linear manner with rising and decreasing slopes, respectively. Increases in macroscopic currents at hyperpolarizing potentials results from a voltage-dependent increase in open probability. Voltage sensitivity is not influenced by divalent cations; however, it is Na+-dependent with a 63-mV decrease in voltage required to reach half-maximal activity per log increase in [Na+]. Mutant channels are particularly sensitive to intracellular [Na+] for removing this sodium abolishes voltage dependence. We conclude that the conserved Trp at the base of TM1 in ENaC/Deg channels protects against voltage by masking an inhibitory allosteric or pore block mechanism, which decreases activity in response to intracellular Na+.
Author List
Pochynyuk O, Kucher V, Boiko N, Mironova E, Staruschenko A, Karpushev AV, Tong Q, Hendron E, Stockand JMESH terms used to index this publication - Major topics in bold
Acid Sensing Ion ChannelsAmino Acid Substitution
Animals
CHO Cells
Cricetinae
Cricetulus
Degenerin Sodium Channels
Epithelial Sodium Channels
Ion Channel Gating
Membrane Potentials
Mice
Mutation, Missense
Nerve Tissue Proteins
Protein Structure, Tertiary
Rats
Sodium Channels
Tryptophan
