Interaction of granulocyte colony-stimulating factor (G-CSF) with its receptor. Evidence that Glu19 of G-CSF interacts with Arg288 of the receptor

Granulocyte colony-stimulating factor (G-CSF) forms a tetrameric complex with its receptor, comprising two G-CSF and two receptor molecules. The structure of the complex is unknown, and it is unclear whether there are one or two binding sites on G-CSF and the receptor. The immunoglobulin-like domain...

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Bibliographic Details
Published in:The Journal of biological chemistry 1999-06, Vol.274 (25), p.17445-17451
Main Authors: Layton, J E, Shimamoto, G, Osslund, T, Hammacher, A, Smith, D K, Treutlein, H R, Boone, T
Format: Article
Language:English
Subjects:
Antigens, CD - genetics
Arginine - chemistry
Binding Sites
Cell Division
Cell Line
Cytokine Receptor gp130
Dimerization
Flow Cytometry
Glutamic Acid - chemistry
Granulocyte Colony-Stimulating Factor - chemistry
Granulocyte Colony-Stimulating Factor - genetics
Humans
Membrane Glycoproteins - genetics
Mutation
Protein Binding
Receptors, Granulocyte Colony-Stimulating Factor - chemistry
Receptors, Granulocyte Colony-Stimulating Factor - genetics
Recombinant Fusion Proteins - genetics
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Summary:Granulocyte colony-stimulating factor (G-CSF) forms a tetrameric complex with its receptor, comprising two G-CSF and two receptor molecules. The structure of the complex is unknown, and it is unclear whether there are one or two binding sites on G-CSF and the receptor. The immunoglobulin-like domain and the cytokine receptor homologous module of the receptor are involved in G-CSF binding, and Arg288 in the cytokine receptor homologous module is particularly important. To identify residues in G-CSF that interact with Arg288, selected charged residues in G-CSF were mutated to Ala. To clarify whether there are two binding sites, a chimeric receptor was created in which the Ig domain was replaced with that of the related receptor gp130. This chimera bound G-CSF but could not transduce a signal, consistent with failure of dimerization and loss of one binding site. The G-CSF mutants had reduced mitogenic activity on cells expressing wild-type receptor. When tested with the chimeric receptor, all G-CSF mutants except one (E46A) showed reduced binding, suggesting that Glu46 is important for interaction with the Ig domain. On cells expressing R288A receptor, all the G-CSF mutants except E19A showed reduced mitogenic activity, indicating that Glu19 of G-CSF interacts with Arg288 of the receptor.
ISSN:0021-9258
1083-351X