Site directed mutagenesis of the complement C5a receptor--examination of a model for its interaction with the ligand C5a.
C5a is a 74 amino acid peptide cleaved from the fifth component of the complement system after activation of either the alternative or classical pathways. It is a potent chemoattractant for neutrophils and monocytes binding to identical receptors on the cell surface. Following the cloning of the cDNA encoding for the human complement C5a receptor, revealing it to be a member of the rhodopsin superfamily of G-protein coupled receptors, a model for the interaction of the C5a receptor with its ligand was proposed, the structure for the receptor being modelled on that of the well defined receptor bacteriorhodopsin. In this model two key residues of the receptor, aspartate82 and either glutamate179 or glutamate 180 were proposed to make up part of the binding site for C5a, acting as counter ions for arginine74 and arginine40, respectively of the C5a molecule. Replacement of aspartate82, glutamate179 and glutamate180 of the C5a receptor with asparagine and glutamine, respectively was shown to have little effect on the dissociation constant of the receptor as detected by Scatchard analysis and competitive binding assays. Hence this modus operandi for the interaction of C5a with its receptor can be rejected.
Krebs Institute, Department of Molecular Biology and Biotechnology, University of Sheffield, U.K.
J E
JE
Pease
D R
DR
Burton
M D
MD
Barker
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D002460:Cell Line; D015936:Complement C5a / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D008956:Models, Chemical; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D044087:Receptor, Anaphylatoxin C5a; D011951:Receptors, Complement / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014169:Transformation, Bacterial
D044087:Receptor, Anaphylatoxin C5a; D011951:Receptors, Complement; D015936:Complement C5a