Elucidation of the CCR1- and CCR5-binding modes of MIP-1α by application of an NMR spectra reconstruction method to the transferred cross-saturation experiments.

C-C chemokine receptor 1 (CCR1) and CCR5 are involved in various inflammation and immune responses, and regulate the progression of the autoimmune diseases differently. However, the number of residues identified at the binding interface was not sufficient to clarify the differences in the CCR1- and CCR5-binding modes to MIP-1α, because the NMR measurement time for CCR1 and CCR5 samples was limited to 24 h, due to their low stability. Here we applied a recently developed NMR spectra reconstruction method, Conservation of experimental data in ANAlysis of FOuRier, to the amide-directed transferred cross-saturation experiments of chemokine receptors, CCR1 and CCR5, embedded in lipid bilayers of the reconstituted high density lipoprotein, and MIP-1α. Our experiments revealed that the residues on the N-loop and β-sheets of MIP-1α are close to both CCR1 and CCR5, and those in the C-terminal helix region are close to CCR5. These results suggest that the genetic influence of the single nucleotide polymorphisms of MIP-1α that accompany substitution of residues in the C-terminal helix region, E57 and V63, would provide clues toward elucidating how the CCR5-MIP-1α interaction affects the progress of autoimmune diseases.