Formation of 1:1 complex of the cytokine receptor homologous region of granulocyte colony-stimulating factor receptor with ligand

The cytokine receptor homologous (CRH) region of the murine granulocyte colony-stimulating factor (G-CSF) receptor was secreted using a Escherichia coli maltose binding protein (MBP) fusion system. The CRH region was prepared from the periplasmic fraction by G-CSF affinity column chromatography and restriction protease factor Xa digestion, and was purified to homogeneity. The purified CRH region specifically bound G-CSF, with an apparent dissociation constant (Kd) of about 1.5 x 10(-9) M. A 1:1 CRH . G-CSF complex was established by gel-filtration high pressure liquid chromatography (HPLC). However, a 2:1 stoichiometric complex was not established, as in the case of the growth hormone (GH) receptor [Recent Prog. Hormone Res., 48, 233-275 (1993)].