Kinetic Resolution of Two Mechanisms for High-Affinity Granulocyte-Macrophage Colony-Stimulating Factor Binding to Its Receptor

Abstract

AbstractGranulocyte-macrophage colony-stimulating factor (GM-CSF) is an important hematopoietic cytokine that exerts its effects by interaction with the GM-CSF receptor (GMR) on the surface of responsive cells. The GM-CSF receptor consists of two subunits: GMRα, which binds GM-CSF with low affinity, and GMRβ, which lacks intrinsic ligand-binding capability but complexes with GMRα to form a high-affinity receptor (GMRα/β). We conducted dynamic kinetic analyses of GM-CSF receptors to define the role of GMRβ in the interaction of ligand and receptor. Our data show that GMRα/β exhibits a higher kon than GMRα, indicating that GMRβ facilitates ligand acquisition to the binding pocket. Heterogeneity with regard to GM-CSF dissociation from GMRα/β points to the presence of loose and tight ligand-receptor complexes in high-affinity binding. Although the loose complex has a koff similar to GMRα, the lower koffindicates that GMRβ inhibits GM-CSF release from the tight receptor complex. The two rates of ligand dissociation may provide for discrete mechanisms of interaction between GM-CSF and its high-affinity receptor. These results show that the β subunit functions to stabilize ligand binding as well as to facilitate ligand acquisition.