UNDERSTANDING AND REDEFNING NEUREGULIN SPECIFICITY

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The highly affinity binding of human neuregulin to the ErbB-3 receptor will be used to analyze the receptor ligand interaction in this family of EGF receptor (ErbB-1) related receptors which is implicated in various forms of human cancer. I will use a combination of computational sequence analysis, directed mutagenesis and phage display selection to construct a function based map of the ligand- receptor interface. This analysis will result in two libraries, each of which represents ten randomized residues on one of the two putative contact surfaces between neuregulin and its receptor. These libraries will be used separately and in combination for the phage display selection of neuregulin isoforms which show specific binding to the ErbB-2 receptor that is highly homologous to ErbB-3. So far, no natural ligand that binds specifically to ErbB-2 has been fully characterized. This receptor is overexpressed on a number of aggressive breast cancer cell lines. Selected ErbB-2-specific ligands can serve as targeting vehicles for "magic bullets" aimed at ErbB-2 overexpressing breast cancer cells. The usefulness of selected ErbB-2 specific isoforms of neurogulin will be demonstrated through fusion with the catalytic and translocation domains of diphtheria toxin, to generate an ErbB-2 specific directed toxin. The sixty amino acid EGF like domain of the neuregulin isoform heregulin beta, that will be used as the starting point for mutagenesis, will also be crystallized for structure determination. This structure will complement the existing NMR structures of the corresponding domain in EGF and neuregulin hrg-alpha as well as aid in the analysis of ligand receptor interaction.