The amino-terminal domain of CCR2 is both necessary and sufficient for high affinity binding of monocyte chemoattractant protein 1. Receptor activation by a pseudo-tethered ligand.
High affinity binding of monocyte chemoattractant protein 1 (MCP-1) requires the presence of the amino-terminal domain of CCR2, the MCP-1 receptor. Here we report that the 35 amino-terminal residues of CCR2, expressed as a membrane-bound fusion protein, bound MCP-1 with an affinity similar to that of the intact, wild-type receptor. Furthermore, the amino-terminal fusion protein enhanced, in trans, agonist-dependent activation of a CCR2 variant that was engineered to lack the high affinity binding sites for MCP-1. Mutation of highly conserved cysteines in the amino-terminal domain and third extracellular loop of CCR2, but not in the fusion protein, resulted in a dramatic loss of MCP-1 binding, suggesting the existence of a critical intramolecular disulfide bond that positions the amino-terminal protein for ligand interaction. These data indicate that the amino-terminal region of CCR2 is both necessary and sufficient for the high affinity binding of MCP-1 and provide the first direct evidence for activation of a chemokine receptor by a pseudo-tethered ligand. In this model, high affinity binding by the relatively short amino-terminal domain of CCR2 serves to tether MCP-1 and enhance low affinity interactions with distal regions of the receptor.
Monteclaro
F S
FS
Gladstone Institute of Cardiovascular Disease, University of California, San Francisco, California 94141-9100, USA.
Charo
I F
IF
D000067956:Adenylyl Cyclase Inhibitors; D018932:Chemokine CCL2 / Q000378:metabolism* / Q000494:pharmacology; D003545:Cysteine / Q000235:genetics; D008958:Models, Molecular; D009154:Mutation; D010446:Peptide Fragments / Q000378:metabolism; D011485:Protein Binding; D054390:Receptors, CCR2; D019707:Receptors, Chemokine; D018121:Receptors, Cytokine / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction
D000067956:Adenylyl Cyclase Inhibitors; D018932:Chemokine CCL2; D010446:Peptide Fragments; D054390:Receptors, CCR2; D019707:Receptors, Chemokine; D018121:Receptors, Cytokine; D011993:Recombinant Fusion Proteins; D003545:Cysteine