Interaction between the C5a receptor and Gi in both the membrane-bound and detergent-solubilized states.

Abstract

C5a elicits a variety of responses from the polymorphonuclear leukocyte all of which utilize G proteins as transducing elements. In the present study, we report the consequences of the interaction between the C5a receptor and the G proteins and describe a system which may allow identification of the transducing proteins. C5a binding to polymorphonuclear leukocyte membranes is inhibited by pertussis, but not cholera, toxin and by a variety of guanine nucleotides. In the absence of nucleotide, we observed a single class of sites with a Kd of 17 pM. The presence of guanosine 5'-3-O-(thio)triphosphate (GTP gamma S) did not alter this affinity but did result in a concentration-dependent decrease in the number of binding sites. Surprisingly, we did not observe the concomitant appearance of a low affinity state implying that, if such a state exists, its affinity is below our limit of detection (5 nM). The receptor and G protein retained their functional interaction following solubilization of the membrane in digitonin. In the absence of nucleotide, we observed a single class of sites with a Kd of 28 pM. Addition of GTP gamma S suppressed binding, and, as was found in membranes, this inhibition is due almost entirely to a decrease in the number of sites. Again we failed to detect the appearance of a lower affinity state. Gel filtration studies of the detergent-solubilized receptor and receptor-C5a complexes indicate that the receptor is precoupled to G protein in the absence of ligand (C5a).

• Authors: S J Siciliano, T E Rollins, M S Springer
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 265 Issue 32 Pg. 19568-74 (Nov 15 1990) ISSN: 0021-9258 [Print] United States
• PMID: 2123189 (Publication Type: Journal Article)

Chemical References

• Guanine Nucleotides
• Receptor, Anaphylatoxin C5a
• Receptors, Complement
• Virulence Factors, Bordetella
• Guanosine 5'-O-(3-Thiotriphosphate)
• Complement C5a
• Cholera Toxin
• GTP-Binding Proteins
• Digitonin

Topics

• Binding Sites (drug effects)
• Cell Membrane (metabolism)
• Cholera Toxin (pharmacology)
• Chromatography, Gel
• Complement C5a (metabolism)
• Digitonin
• GTP-Binding Proteins (metabolism)
• Guanine Nucleotides (pharmacology)
• Guanosine 5'-O-(3-Thiotriphosphate) (pharmacology)
• Humans
• Neutrophils (metabolism)
• Receptor, Anaphylatoxin C5a
• Receptors, Complement (isolation & purification, metabolism)
• Signal Transduction
• Temperature
• Virulence Factors, Bordetella (pharmacology)