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 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.
Layton
J E
JE
Ludwig Institute for Cancer Research, Melbourne Tumour Biology Branch, Parkville 3050, Australia. Judy.Layton@ludwig.edu.au
Shimamoto
G
G
Osslund
T
T
Hammacher
A
A
Smith
D K
DK
Treutlein
H R
HR
Boone
T
T
D015703:Antigens, CD / Q000235:genetics; D001120:Arginine / Q000737:chemistry*; D001665:Binding Sites; D002455:Cell Division; D002460:Cell Line; D050822:Cytokine Receptor gp130; D019281:Dimerization; D005434:Flow Cytometry; D018698:Glutamic Acid / Q000737:chemistry*; D016179:Granulocyte Colony-Stimulating Factor / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics; D009154:Mutation; D011485:Protein Binding; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000737:chemistry* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics
D015703:Antigens, CD; C494199:IL6ST protein, human; D008562:Membrane Glycoproteins; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D011993:Recombinant Fusion Proteins; D050822:Cytokine Receptor gp130; D016179:Granulocyte Colony-Stimulating Factor; D018698:Glutamic Acid; D001120:Arginine