Chemokines are small soluble proteins that stimulate leukocyte recruitment by activation of chemokine receptors, a subfamily of G protein-coupled receptors (GPCRs) expressed on leukocytes. Chemokine-receptor interactions play critical roles in a wide range of inflammatory diseases, such as multiple sclerosis and atherosclerosis, as well as in HIV, cancer and diabetes.

Among the chemokine receptor family, CCR1 is particularly interesting, as it is already recognised as a drug target for multiple sclerosis and rheumatoid arthritis; some small non-peptide CCR1 antagonists have been reported. However, the structural basis of chemokine recognition by CCR1 is largely unexplored. Notably, CCR1 contains two tyrosine residues in its extracellular N-terminal region that are potential sulfation sites, which presents a high interest as sulfation has already been shown as an important post-translational modification for chemokine receptor signalling. CCR1 recognises more than eight CC chemokines, including RANTES/CCL5, HCC-1/CCL14, HCC-2/CCL15 and MPIF-1/CCL23.

To characterise the interactions between the N-terminal region of CCR1 and CCR1-cognate chemokines, we have studied binding between four sulfopeptides, corresponding to different sulfation states of the CCR1 N-terminus, with several chemokines. Sulfopeptide were obtained by solid phase peptide synthesis, whereas chemokines were expressed in E. coli inclusion bodies, folded and purified. Binding affinities were determined using fluorescence anisotropy assays, thus indicating the influence of tyrosine sulfation on chemokine binding and selectivity. NMR spectroscopy is being used to characterise the structural basis of sulfotyrosine recognition. This study will help us to understand the role of tyrosine sulfation in chemokine-chemokine receptor interactions.