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Stabilization of C5a receptor–G‐protein interactions through ligand binding
Author(s) -
Wennogle Lawrence P.,
Conder Lynnette,
Winter Cindy,
Braunwalder Albert,
Vlattas Sid,
Kramer Richard,
Cioffi Catherine,
Hu ShouIh
Publication year - 1994
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.240550316
Subject(s) - c5a receptor , receptor , 5 ht5a receptor , affinity chromatography , protease activated receptor 2 , biotinylation , biochemistry , biology , microbiology and biotechnology , chemistry , antibody , enzyme , complement system , immunology
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 i alpha 2 and G i alpha 3 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.