Attempts to define functional domains of gap junction proteins with synthetic peptides.
To map the binding sites involved in channel formation, synthetic peptides representing sequences of connexin 32 were tested for their ability to inhibit cell-cell channel formation. Both large peptides representing most of the two presumed extracellular loops of connexin32 and shorter peptides representing subsets of these larger peptides were found to inhibit cell-cell channel formation. The properties of the peptide inhibition suggested that the binding site is complex, involving several segments of both extracellular loops. One of the peptides (a 12-mer) did not inhibit but instead was found to form channels in membranes. Both in oocyte membranes and in bilayers, the channels formed by the peptide were asymmetrically voltage dependent. Their unit conductances ranged from 20 to 160 pS. These data are discussed in the form of a model in which the connexin sequence represented by the peptide is part of a beta structure providing the lining of the channel pore.
Department of Physiology and Biophysics, University of Miami, School of Medicine, Florida 33101.
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Dahl
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Nonner
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Werner
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D055592:Biophysical Phenomena; D001703:Biophysics; D017630:Connexins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D004285:Dogs; D005260:Female; D066298:In Vitro Techniques; D008051:Lipid Bilayers; D008956:Models, Chemical; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000378:metabolism; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology*; D010449:Peptide Mapping; D012333:RNA, Messenger / Q000235:genetics; D011817:Rabbits; D014982:Xenopus laevis; D000097002:Gap Junction beta-1 Protein
D017630:Connexins; D008051:Lipid Bilayers; D010446:Peptide Fragments; D012333:RNA, Messenger; D000097002:Gap Junction beta-1 Protein