Stabilization of C5a receptor–G-protein interactions through ligand binding

Binding of biotin-C5a to the C5a receptor in membrane fragments followed by detergent solubilization and purification with streptavidin-agarose affinity chromatography resulted in the isolation of a receptor complex with associated G-proteins. In contrast, when receptor was detergent-solubilized in the absence of C5a and purified by affinity chromatography with Affigel-C5a, G-proteins did not copurify. Since the results indicate that receptor ligation stabilized the receptor–G-protein interaction to allow purification of the complex, the findings emphasize the dynamic nature of the C5a receptor–effector interactions. When biotin-C5a–ligated receptor was purified from a mouse cell line overexpressing recombinant human receptor, both G_ialpha₂ and G_ialpha₃ subunits copurified, confirming that multiple transducing systems are linked to the C5a receptor. The method of stabilization of receptor-transducer complexes offers the opportunity to further elaborate the interactions of the C5a receptor with diverse transducing elements and second messenger systems. © 1994 Wiley-Liss, Inc.