Evidence for the ligand-induced conversion from a dimer to a tetramer of the granulocyte colony-stimulating factor receptor.
An extracellular portion of granulocyte colony-stimulating factor (G-CSF) receptor, which contains an immunoglobulin-like (Ig) domain and cytokine receptor homologous (CRH) region, was secreted into the medium using Trichoplusia ni-Autographa californica nuclear polyhedrosis virus system. The gene product was purified to homogeneity mainly as a dimer (85 kDa) using G-CSF affinity column chromatography and gel filtration HPLC, although the product existed as a monomer (45 kDa) in the medium. Scatchard analyses suggested that only the dimer had high affinity ligand binding (Kd = about 100 pM), which is comparable with the Kd value of the cell surface receptor. The binding of G-CSF to Ig-CRH induced its tetramerization (200-250 kDa). The molecular composition of the tetrameric complex showed a stoichiometry of four ligands bound to four Ig-CRH. These results suggested that the oligomeric mechanism of the G-CSF receptor differs from that reported for growth hormone (GH) receptor, although CD spectrum spectroscopy suggested that the Ig-CRH has a GH receptor-like structure.
Hiraoka
O
O
Protein Engineering Research Institute, Osaka, Japan.
Anaguchi
H
H
Ota
Y
Y
D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D017931:DNA Primers; D005822:Genetic Vectors; D007313:Insecta; D008024:Ligands; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D017924:Nucleopolyhedroviruses; D016133:Polymerase Chain Reaction; D018121:Receptors, Cytokine / Q000737:chemistry; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D017931:DNA Primers; D008024:Ligands; D046911:Macromolecular Substances; D018121:Receptors, Cytokine; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D011994:Recombinant Proteins