Purification and characterization of the receptor for murine granulocyte colony-stimulating factor.

A receptor for mouse granulocyte colony-stimulating factor (G-CSF) has been found on the cell surface of mouse myeloid leukemia cell line NFS-60. Chemical cross-linking of the receptor with radioiodinated G-CSF, followed by gel electrophoresis in the presence of sodium dodecyl sulfate, has revealed that the G-CSF receptor in the NFS-60 cells is a single polypeptide of Mr approximately 100,000-130,000. The receptor in the membrane fraction of NFS-60 cells were solubilized in an active form with 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonic acid. The solubilized receptor was purified approximately 100,000-fold to near homogeneity using a G-CSF affinity gel and gel filtration on a Superose 12 column, as measured by the selective precipitation of the 125I-G-CSF-receptor complex by polyethylene glycol. The purified G-CSF receptor has two classes of binding characteristics, one with an equilibrium dissociation constant (Kd) of 120-360 pM which is comparable with the Kd value for the cell-surface receptor, and the other with a higher Kd value of 2.6-4.2 nM. Analyses of the purified receptor by ligand blotting and sucrose density gradient centrifugation indicated that the low-affinity receptor is the monomer of the Mr 100,000-130,000 protein, whereas the high-affinity receptor consists of oligomers of the protein.