title: An inhibitory peptide derived from the α-subunit of the epithelial sodium channel (ENaC) shows a helical conformation.
abstract: Proteolytic activation of the heteromeric epithelial sodium channel (ENaC) is thought to involve the release of inhibitory peptides from the extracellular domains of its α- and γ-subunit. Recently, we demonstrated that an α-13-mer peptide, corresponding to a putative inhibitory region within the extracellular domain of human αENaC, inhibits human αβγENaC. The aim of the present study was to investigate the structural basis of the inhibitory effect of this α-13-mer peptide. Analysis of the peptide by replica exchange molecular dynamics method, circular dichroism spectroscopy, nuclear magnetic resonance spectroscopy, and molecular dynamics simulations suggested that a helical turn at the carboxy-terminus is the preferred conformational state of the α-13-mer peptide. From this we predicted that a specific mutation (leucine 188 to alanine) should have a strong effect on the conformational preferences of the peptide. To functionally test this, we compared the effect of the wild-type α-13-mer with that of a mutant α-L188A-13-mer on ENaC currents in Xenopus laevis oocytes heterologously expressing human αβγENaC. We demonstrated that replacing the leucine 188 by alanine abolished the inhibitory effect of the α-13-mer peptide on ENaC. These findings suggest that a helical conformation in its carboxyterminal part is functionally important to mediate ENaC inhibition by the α-13-mer peptide. However, high resolution structural information on the complex of the inhibitory αENaC peptide and the channel are needed to confirm this conclusion.
authors.affiliation: Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Silke
authors.initials: S
authors.lastname: Haerteis
authors.forename: Daniel
authors.initials: D
authors.lastname: Schaal
authors.forename: Felix
authors.initials: F
authors.lastname: Brauer
authors.forename: Stefan
authors.initials: S
authors.lastname: Brüschke
authors.forename: Kristian
authors.initials: K
authors.lastname: Schweimer
authors.forename: Robert
authors.initials: R
authors.lastname: Rauh
authors.forename: Heinrich
authors.initials: H
authors.lastname: Sticht
authors.forename: Paul
authors.initials: P
authors.lastname: Rösch
authors.forename: Stephan
authors.initials: S
authors.lastname: Schwarzinger
authors.forename: Christoph
authors.initials: C
authors.lastname: Korbmacher
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D053503:Epithelial Sodium Channels / Q000737:chemistry*; D006801:Humans; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry*; D011487:Protein Conformation; D014982:Xenopus laevis
chemical_list: D053503:Epithelial Sodium Channels; D010446:Peptide Fragments