title: Solution NMR characterization of chemokine CXCL8/IL-8 monomer and dimer binding to glycosaminoglycans: structural plasticity mediates differential binding interactions.
abstract: Chemokine CXCL8/interleukin-8 (IL-8) plays a crucial role in directing neutrophils and oligodendrocytes to combat infection/injury and tumour cells in metastasis development. CXCL8 exists as monomers and dimers and interaction of both forms with glycosaminoglycans (GAGs) mediate these diverse cellular processes. However, very little is known regarding the structural basis underlying CXCL8-GAG interactions. There are conflicting reports on the affinities, geometry and whether the monomer or dimer is the high-affinity GAG ligand. To resolve these issues, we characterized the binding of a series of heparin-derived oligosaccharides [heparin disaccharide (dp2), heparin tetrasaccharide (dp4), heparin octasaccharide (dp8) and heparin 14-mer (dp14)] to the wild-type (WT) dimer and a designed monomer using solution NMR spectroscopy. The pattern and extent of binding-induced chemical shift perturbation (CSP) varied between dimer and monomer and between longer and shorter oligosaccharides. NMR-based structural models show that different interaction modes coexist and that the nature of interactions varied between monomer and dimer and oligosaccharide length. MD simulations indicate that the binding interface is structurally plastic and provided residue-specific details of the dynamic nature of the binding interface. Binding studies carried out under conditions at which WT CXCL8 exists as monomers and dimers provide unambiguous evidence that the dimer is the high-affinity GAG ligand. Together, our data indicate that a set of core residues function as the major recognition/binding site, a set of peripheral residues define the various binding geometries and that the structural plasticity of the binding interface allows multiplicity of binding interactions. We conclude that structural plasticity most probably regulates in vivo CXCL8 monomer/dimer-GAG interactions and function.
authors.affiliation: Department of Biochemistry and Molecular Biology, The University of Texas Medical Branch, Galveston, TX 77555, U.S.A. Sealy Center for Structural Biology and Molecular Biophysics, The University of Texas Medical Branch, Galveston, TX 77555, U.S.A. Institute for Structural Biology, Drug Discovery and Development, Virginia Commonwealth University, Richmond, VA 23219, U.S.A.
authors.forename: Prem Raj B
authors.initials: PR
authors.lastname: Joseph
authors.affiliation: Department of Medicinal Chemistry, Virginia Commonwealth University, Richmond, VA 23219, U.S.A. Institute for Structural Biology, Drug Discovery and Development, Virginia Commonwealth University, Richmond, VA 23219, U.S.A.
authors.forename: Philip D
authors.initials: PD
authors.lastname: Mosier
authors.affiliation: Department of Medicinal Chemistry, Virginia Commonwealth University, Richmond, VA 23219, U.S.A.
authors.forename: Umesh R
authors.initials: UR
authors.lastname: Desai
authors.affiliation: Department of Biochemistry and Molecular Biology, The University of Texas Medical Branch, Galveston, TX 77555, U.S.A. Sealy Center for Structural Biology and Molecular Biophysics, The University of Texas Medical Branch, Galveston, TX 77555, U.S.A. krrajara@utmb.edu.
authors.forename: Krishna
authors.initials: K
authors.lastname: Rajarathnam
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D006025:Glycosaminoglycans / Q000737:chemistry* / Q000378:metabolism; D006493:Heparin / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D016209:Interleukin-8 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D012996:Solutions
chemical_list: C508606:CXCL8 protein, human; D006025:Glycosaminoglycans; D016209:Interleukin-8; D009844:Oligosaccharides; D012996:Solutions; D006493:Heparin
keywords: chemokine; glycosaminoglycan (GAG); heparin; nuclear magnetic resonance (NMR); structural plasticity; structure–function study