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 Gialpha2 and Gialpha3 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.
Research Department, CIBA-GEIGY Pharmaceuticals Division, Summit, New Jersey 07901.
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Wennogle
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Conder
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Winter
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Braunwalder
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Vlattas
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Kramer
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Cioffi
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D000818:Animals; D002460:Cell Line; D015936:Complement C5a / Q000302:isolation & purification / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D008024:Ligands; D051379:Mice; D044087:Receptor, Anaphylatoxin C5a; D011951:Receptors, Complement / Q000302:isolation & purification / Q000378:metabolism*
D008024:Ligands; D044087:Receptor, Anaphylatoxin C5a; D011951:Receptors, Complement; D015936:Complement C5a; D019204:GTP-Binding Proteins