title: Ligand binding characteristics of the carboxyl-terminal domain of the cytokine receptor homologous region of the granulocyte colony-stimulating factor receptor.
abstract: The carboxyl-terminal domain (BC domain, roughly 100 amino acid residues) of the cytokine receptor homologous region in the receptor for murine granulocyte colony-stimulating factor was secreted as a maltose binding protein fusion into the Escherichia coli periplasm. The murine BC domain was prepared from the fusion protein by restriction protease factor Xa digestion and was purified to homogeneity. The purified BC domain specifically and stoichiometrically bound granulocyte colony-stimulating factor. This result indicates that the BC domain is also critical for ligand binding, as shown for the amino-terminal domain of the cytokine receptor homologous region (Hiraoka, O., Anaguchi, H., Yamasaki, K., Fukunaga, R., Nagata, S., and Ota, Y. (1994) J. Biol. Chem. 269, 22412-22419). The tertiary folding and the beta-sheet structure of the BC domain were confirmed by NMR spectroscopy. The disulfide bond pattern suggested from peptide mapping was Cys224-Cys271 and Cys242-Cys285. Disruption of the disulfide bonds suggested that both bonds are critical for maintaining the folding of the BC domain, although a BC domain lacking the second bond still retained ligand binding activity. Mutational analysis of the WSXWS sequence conserved in the cytokine receptor family suggested that this motif is critical for protein folding rather than for ligand binding.
mesh_terms: D018528:ATP-Binding Cassette Transporters; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000096:biosynthesis; D003001:Cloning, Molecular; D004220:Disulfides / Q000032:analysis; D004926:Escherichia coli; D029968:Escherichia coli Proteins; D015951:Factor Xa / Q000378:metabolism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D058619:Maltose-Binding Proteins; D051379:Mice; D008961:Models, Structural; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010449:Peptide Mapping; D010957:Plasmids; D017433:Protein Structure, Secondary; D018121:Receptors, Cytokine / Q000737:chemistry*; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D002352:Carrier Proteins; D004220:Disulfides; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments; D018121:Receptors, Cytokine; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D011993:Recombinant Fusion Proteins; C461599:maltose transport system, E coli; D015951:Factor Xa