title: Structure of the amino terminus of a gap junction protein.
abstract: Charged amino acid residues in the amino terminus of gap junction forming proteins (connexins) form part, if not all, of the transjunctional voltage sensor of gap junction channels and play a fundamental role in ion permeation. Results from studies of the voltage dependence of N-terminal mutants predict that residues 1-10 of Group I connexins lie within the channel pore and that the N-terminus forms the channel vestibule by the creation of a turn initiated by the conserved G12 residue. Here we report that intercellular channels containing mutations of G12 in Cx32 to residues that are likely to interfere with flexibility of this locus (G12S, G12Y, and G12V) do not express junctional currents, whereas a connexin containing a proline residue at G12 (Cx32G12P), which is expected to maintain a structure similar to that of the G12 locus, forms nearly wild-type channels. We have solved the structure of an N-terminal peptide of Cx26 (MDWGTLQSILGGVNK) using 1H 2D NMR. The peptide contains two structured domains connected by a flexible hinge (domain-hinge-domain motif) that would allow the placement of the amino terminus within the channel pore. Residues 1-10 adopt a helical conformation and line the channel entrance while residues 12-15 form an open turn. Overall, there is good agreement between the structural and dynamic features of the N-terminal peptide provided by NMR and the functional studies of the voltage dependence of channels formed by wild-type and N-terminal mutations.
authors.affiliation: Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
authors.forename: P E
authors.initials: PE
authors.lastname: Purnick
authors.forename: D C
authors.initials: DC
authors.lastname: Benjamin
authors.forename: V K
authors.initials: VK
authors.lastname: Verselis
authors.forename: T A
authors.initials: TA
authors.lastname: Bargiello
authors.forename: T L
authors.initials: TL
authors.lastname: Dowd
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000072259:Connexin 26; D017630:Connexins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017629:Gap Junctions / Q000737:chemistry; D006801:Humans; D066298:In Vitro Techniques; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019906:Nuclear Magnetic Resonance, Biomolecular; D009865:Oocytes / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D055672:Static Electricity; D013816:Thermodynamics; D014981:Xenopus
chemical_list: D017630:Connexins; C000607045:GJB2 protein, Xenopus; C000607042:GJB2 protein, human; D010446:Peptide Fragments; D011994:Recombinant Proteins; D000072259:Connexin 26