TY - JOUR

T1 - Positive versus negative modulation of different endogenous chemokines for CC-chemokine receptor 1 by small molecule agonists through allosteric versus orthosteric binding

AU - Jensen, Pia C

AU - Thiele, Stefanie

AU - Ulven, Trond

AU - Schwartz, Thue W

AU - Rosenkilde, Mette M

PY - 2008/6/17

Y1 - 2008/6/17

N2 - 7 transmembrane-spanning (7TM) chemokine receptors having multiple endogenous ligands offer special opportunities to understand the molecular basis for allosteric mechanisms. Thus, CC-chemokine receptor 1 (CCR1) binds CC-chemokine 3 and 5 (CCL3 and CCL5) with Kd values of 7.3 and 0.16 nM, respectively, as determined in homologous competition binding assays. However, CCL5 appears to have a >10.000 fold lower affinity in competition against 125I-CCL3. Mutational mapping revealed that CCL3 and CCL5 both are strongly affected by systematic truncations of the N-terminal extension, whereas only CCL5 and not CCL3 activation is affected by substitutions in the main ligand binding pocket including the conserved GluVII:06 anchor point. A series of metal-ion chelator complexes were found to act as full agonists on CCR1 and to be critically affected by the same substitutions in the main ligand binding pocket as CCL5 but not by mutations in the extracellular domain. In agreement with the overlapping binding sites, the small non-peptide agonists displaced radiolabeled CCL5 with high affinity. Interestingly, the same compounds acted as allosteric enhancers of the binding of CCL3 - with which they did not overlap in binding site - leading to an increased Bmax and affinity of this chemokine mainly due to an increased association rate. It is concluded that a small molecule agonist through binding deep in the main ligand binding pocket can act as an allosteric enhancer for one endogenous chemokine and at the same time as a competitive blocker of the binding of another endogenous chemokine.

AB - 7 transmembrane-spanning (7TM) chemokine receptors having multiple endogenous ligands offer special opportunities to understand the molecular basis for allosteric mechanisms. Thus, CC-chemokine receptor 1 (CCR1) binds CC-chemokine 3 and 5 (CCL3 and CCL5) with Kd values of 7.3 and 0.16 nM, respectively, as determined in homologous competition binding assays. However, CCL5 appears to have a >10.000 fold lower affinity in competition against 125I-CCL3. Mutational mapping revealed that CCL3 and CCL5 both are strongly affected by systematic truncations of the N-terminal extension, whereas only CCL5 and not CCL3 activation is affected by substitutions in the main ligand binding pocket including the conserved GluVII:06 anchor point. A series of metal-ion chelator complexes were found to act as full agonists on CCR1 and to be critically affected by the same substitutions in the main ligand binding pocket as CCL5 but not by mutations in the extracellular domain. In agreement with the overlapping binding sites, the small non-peptide agonists displaced radiolabeled CCL5 with high affinity. Interestingly, the same compounds acted as allosteric enhancers of the binding of CCL3 - with which they did not overlap in binding site - leading to an increased Bmax and affinity of this chemokine mainly due to an increased association rate. It is concluded that a small molecule agonist through binding deep in the main ligand binding pocket can act as an allosteric enhancer for one endogenous chemokine and at the same time as a competitive blocker of the binding of another endogenous chemokine.

U2 - 10.1074/jbc.M803458200

DO - 10.1074/jbc.M803458200

M3 - Journal article

VL - 283

SP - 23121

EP - 23128

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 34

ER -