Hydrophobic interactions mediate binding of the glycine receptor beta-subunit to gephyrin.
Glycine receptors (GlyRs) are ligand-gated chloride channel proteins composed of alpha- and beta-subunits. GlyRs are located to and anchored at postsynaptic sites by the receptor-associated protein gephyrin. Previous work from our laboratory has identified a core motif for gephyrin binding in the cytoplasmic loop of the GlyR beta-subunit. Here, we localized amino acid residues implicated in gephyrin binding by site-directed mutagenesis. In a novel transfection assay, a green fluorescent protein-gephyrin binding motif fusion protein was used to monitor the consequences of amino acid substitutions for beta-subunit interaction with gephyrin. Only multiple, but not single, replacements of hydrophobic side chains abolished the interaction between the two proteins. Our data are consistent with gephyrin binding being mediated by the hydrophobic side of an imperfect amphipathic helix.
Kneussel
M
M
Department of Neurochemistry, Max-Planck-Institute for Brain Research, Frankfurt/Main, Germany.
Hermann
A
A
Kirsch
J
J
Betz
H
H
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D002460:Cell Line; D049452:Green Fluorescent Proteins; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D018009:Receptors, Glycine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D008164:Luminescent Proteins; D008565:Membrane Proteins; D018009:Receptors, Glycine; D011993:Recombinant Fusion Proteins; C073441:gephyrin; D049452:Green Fluorescent Proteins