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Identification of a ligand-binding site on the granulocyte colony-stimulating factor receptor by molecular modeling and mutagenesis.
- Source :
-
The Journal of biological chemistry [J Biol Chem] 1997 Nov 21; Vol. 272 (47), pp. 29735-41. - Publication Year :
- 1997
-
Abstract
- Granulocyte colony-stimulating factor (G-CSF) initiates its effects on cells of the neutrophil lineage by inducing formation of a homodimeric receptor complex. The structure of the G-CSF receptor has not yet been determined, therefore we used molecular modeling to identify regions of the receptor that were likely to be involved in ligand binding. The G-CSF receptor sequence was aligned with all the available sequences of the gp130 and growth hormone receptor families and a model of the cytokine receptor homologous domain was constructed, based on the growth hormone receptor structure. Alanine substitution mutagenesis was performed on loops and individual residues that were predicted to bind ligand. Mutant receptors were expressed in factor-dependent Ba/F3 cells and assessed for proliferation response and ligand binding. Six residues were identified that significantly reduced receptor function, with Arg288 in the F'-G' loop having the greatest effect. These residues formed a binding face on the receptor model resembling the growth hormone receptor site, which suggests that the model is reasonable. However, electrostatic analysis of the model provided further evidence that the mechanism of receptor dimerization is different from that of the growth hormone receptor.
- Subjects :
- Amino Acid Sequence
Animals
Binding Sites
Corticotropin-Releasing Hormone metabolism
Flow Cytometry
Ligands
Mice
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Receptors, Granulocyte Colony-Stimulating Factor chemistry
Receptors, Granulocyte Colony-Stimulating Factor genetics
Sequence Alignment
Static Electricity
Tumor Cells, Cultured
Receptors, Granulocyte Colony-Stimulating Factor metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0021-9258
- Volume :
- 272
- Issue :
- 47
- Database :
- MEDLINE
- Journal :
- The Journal of biological chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 9368043
- Full Text :
- https://doi.org/10.1074/jbc.272.47.29735