title: Use of a modified bacteriophage to probe the interactions between peptides and ion channel receptors in mammalian cells.
abstract: Besides natural peptide ligands, screening of random peptide libraries has yielded novel bioactive peptides for cell surface receptors. A method is described that uses a modified bacteriophage as a detection reagent to monitor the expression of receptor channels in mammalian cells and to probe the molecular interaction between phage-tethered peptides (phi T-peptides) and specific receptor targets. By taking advantage of a specific multivalent interaction between phi T-peptides and the receptor target, assays have been developed that use phi T-peptides specific for the N-methyl-D-aspartate glutamate receptor, an important ligand-gated ion channel in the nervous system, to monitor the receptor expression in cultured mammalian cells. Combining these phi T-peptide binding assays with fluorescence-activated cell sorting, 10(4) random glutamate receptor mutants were screened and candidate interaction residues were identified. This dual heterologous expression system offers a powerful approach to the molecular studies of protein-protein interactions.
authors.affiliation: Department of Physiology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. min-li@qmail.bs.jhu.edu
authors.forename: M
authors.initials: M
authors.lastname: Li
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001435:Bacteriophages; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D005434:Flow Cytometry; D008322:Mammals; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019151:Peptide Library; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D017470:Receptors, Glutamate / Q000737:chemistry* / Q000378:metabolism*; D016194:Receptors, N-Methyl-D-Aspartate / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D016324:Sequence Tagged Sites
chemical_list: D017931:DNA Primers; D019151:Peptide Library; D010455:Peptides; D017470:Receptors, Glutamate; D016194:Receptors, N-Methyl-D-Aspartate; D011994:Recombinant Proteins