TI  - Mutagenesis studies of the human <prot><prot>erythropoietin</prot> receptor</prot>. Establishment of
      structure-function relationships.
PG  - 4985-92
AB  - Mutagenesis of the  <prot><prot>erythropoietin</prot> receptor</prot> (<prot>EPOR</prot>)  permits analysis of the
      contribution that individual amino acid residues make to <prot>erythropoietin</prot>
       (<prot>EPO</prot>)  binding. We employed both random and site-specific mutagenesis to
      determine the function of amino acid residues in the extracellular domain
      (referred to as <prot><prot>EPO</prot> binding protein</prot>, <prot>EBP</prot>) of the <prot>EPOR</prot>. Residues were
      chosen for site-specific alanine substitution based on the results of the
      random mutagenesis or on their homology to residues that are conserved or
      have been reported to be involved in ligand binding in other receptors of
      the cytokine receptor family. Site-specific mutants were expressed in
      Escherichia coli as soluble <prot>EBP</prot> and analyzed for <prot>EPO</prot> binding in several
      different assay formats. In addition, selected mutant proteins were
      expressed as full-length <prot>EPOR</prot> on the surface of COS cells and analyzed for
      125I-<prot>EPO</prot> binding in receptor binding assays. Using these methods, we have
      identified residues that appear to be involved in  <prot>EPO</prot>  binding as well as
      other residues, most of which are conserved in receptors of the cytokine
      receptor family, that appear to be necessary for the proper folding and/or
      stability of the  <prot>EPOR</prot>.  We present correlations between these mutagenesis
      data and the recently solved crystal structure of the <prot>EBP</prot> with a peptide
      ligand.
AD  - The R. W. Johnson Pharmaceutical Research Institute, Drug Discovery
