title: Exploration of the pore structure of a peptide-gated Na+ channel.
abstract: The FMRF-amide-activated sodium channel (FaNaC), a member of the ENaC/Degenerin family, is a homotetramer, each subunit containing two transmembrane segments. We changed independently every residue of the first transmembrane segment (TM1) into a cysteine and tested each position's accessibility to the cysteine covalent reagents MTSET and MTSES. Eleven mutants were accessible to the cationic MTSET, showing that TM1 faces the ion translocation pathway. This was confirmed by the accessibility of cysteines present in the acid-sensing ion channels and other mutations introduced in FaNaC TM1. Modification of accessibilities for positions 69, 71 and 72 in the open state shows that the gating mechanism consists of the opening of a constriction close to the intracellular side. The anionic MTSES did not penetrate into the channel, indicating the presence of a charge selectivity filter in the outer vestibule. Furthermore, amiloride inhibition resulted in the channel occlusion in the middle of the pore. Summarizing, the ionic pore of FaNaC includes a large aqueous cavity, with a charge selectivity filter in the outer vestibule and the gate close to the interior.
authors.affiliation: Laboratoire de Physiologie Cellulaire et Moléculaire, CNRS UMR6548, Université de Nice-Sophia Antipolis, Faculté des Sciences, Parc Valrose, 06108 Nice, France.
authors.forename: M
authors.initials: M
authors.lastname: Poët
authors.forename: M
authors.initials: M
authors.lastname: Tauc
authors.forename: E
authors.initials: E
authors.lastname: Lingueglia
authors.forename: P
authors.initials: P
authors.lastname: Cance
authors.forename: P
authors.initials: P
authors.lastname: Poujeol
authors.forename: M
authors.initials: M
authors.lastname: Lazdunski
authors.forename: L
authors.initials: L
authors.lastname: Counillon
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D003545:Cysteine / Q000737:chemistry; D018076:DNA, Complementary / Q000235:genetics; D019835:FMRFamide / Q000494:pharmacology*; D005260:Female; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects*; D017136:Ion Transport / Q000187:drug effects*; D008698:Mesylates / Q000494:pharmacology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D005810:Multigene Family; D016297:Mutagenesis, Site-Directed; D009865:Oocytes; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012964:Sodium / Q000378:metabolism*; D015222:Sodium Channels / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D055672:Static Electricity; D013439:Sulfhydryl Reagents / Q000494:pharmacology; D014982:Xenopus laevis
chemical_list: D018076:DNA, Complementary; C097095:FaNaCh protein, Helix aspersa; D008698:Mesylates; D011993:Recombinant Fusion Proteins; D015222:Sodium Channels; D013439:Sulfhydryl Reagents; C109153:(2-sulfonatoethyl)methanethiosulfonate; C109154:(2-(trimethylammonium)ethyl)methanethiosulfonate; D019835:FMRFamide; D012964:Sodium; D003545:Cysteine