Solution structure and dynamics of the CX3C chemokine domain of fractalkine and its interaction with an N-terminal fragment of CX(3)CR1

Fractalkine, a novel CX3C chemokine, is unusual because of both its membran e-associated structure and its direct role in cell adhesion. We have solved the solution structure of the chemokine domain of fractalkine (residues 1- 76) by heteronuclear NMR methods. The 20 lowest energy structures in the en semble have an average backbone rmsd of 0.43 Angstrom, excluding the termin i. In contrast to many other chemokines which form homodimers, fractalkine' s chemokine module is monomeric. Comparison of the structure to CC and CXC chemokines reveals interesting differences which are Likely to be relevant to receptor binding. These include a bulge formed by the CX3C motif, the re lative orientation of the N-terminus and 30's loop (residues 30-38), and th e conformation of the N-loop (residues 9-19). N-15 backbone relaxation expe riments indicate that these same regions of the protein are dynamic. We als o titrated N-15-labeled protein with a peptide from the N-terminus of the r eceptor CX(3)CR1 and confirmed that this region of the receptor contacts th e fractalkine chemokine domain. Interestingly, the binding site maps roughl y to the regions of greatest flexibility and structural variability. Togeth er, these data provide a first glimpse of how fractalkine interacts with it s receptor and should help guide mutagenesis studies to further elucidate t he molecular details of binding and signaling through CX(3)CR1.