“Glycosaminoglycan Disaccharide Alters the Dimer Dissociation Constant of the Chemokine MIP-1b” Melissa A. McCornack, Danielle M. Boren, and Patricia J. LiWang Biochemistry 43 10090-10101 (2004).

Binding Heparin disaccharide significantly tightens the MIP-1b dimer:

Chemokines are immune system proteins that recruit and activate leukocytes to sites of infection. This recruitment is believed to involve the establishment of a chemokine concentration gradient by the binding of chemokines to glycosaminoglycans (GAGs). In previous studies, we elucidated the GAG binding site of the chemokine MIP-1b and implicated the involvement of the chemokine dimer in GAG binding through residues across the dimer interface. In the present studies, nuclear magnetic resonance spectroscopy was used to investigate the effect of GAG binding on MIP-1b dimerization. Using several dimerization-impaired variants of MIP-1b (F13Y, F13L, L34W, and L34K), these studies indicate that the addition of disaccharide to the mutants increases their dimerization affinities. For MIP-1b F13Y, the presence of the disaccharide increases the chemokine dimerization affinity about nine fold as evidenced by a decrease in the dimer dissociation constant from 610 mM to 66 mM. Even more dramatically, the dimerization affinity of MIP-1b L34W also increases upon addition of disaccharide with the dimer dissociation constant decreasing from 97 mM to 6.5 mM. Having shown this effect for the mutants of MIP-1b, similar experiments were conducted with the CC chemokine RANTES, and it was demonstrated that the presence of disaccharide increases its dimerization affinity by almost seven fold. These findings provide further evidence of the importance of the dimer in chemokine function and provide the first quantitative investigation of the role of GAGs in the manipulation of MIP-1b quaternary structure.