title: Receptor engagement by viral interleukin-6 encoded by Kaposi sarcoma-associated herpesvirus.
abstract: Receptor usage by viral interleukin-6 (vIL-6), a virokine encoded by Kaposi sarcoma- associated herpesvirus, is an issue of controversy. Recently, the crystal structure of vIL-6 identified vIL-6 sites II and III as directly binding to glycoprotein (gp)130, the common signal transducer for the IL-6 family of cytokines. Site I of vIL-6, however, comprising the outward helical face of vIL-6, where human IL-6 (hIL-6) would interact with the specific alpha-chain IL-6 receptor (IL-6R), is accessible and not occupied by gp130. This study examined whether this unused vIL-6 surface is available for IL-6R binding. By enzyme-linked immunosorbent assay, vIL-6 bound to soluble gp130 (sgp130) but not to soluble IL-6R (sIL-6R). Using plasmon surface resonance, vIL-6 bound to sgp130 with a dissociation constant of 2.5 microM, corresponding to 1000-fold lower affinity than that of hIL-6/sIL-6R complex for gp130. sIL-6R neither bound to vIL-6 nor affected vIL-6 binding to gp130. In bioassays, vIL-6 activity was neutralized by 4 monoclonal antibodies (mAbs) recognizing a domain within vIL-6 site I, mapped to the C-terminal part of the AB-loop and the beginning of helix B. The homologous region in hIL-6 participates in site I binding to IL-6R. In addition, binding of vIL-6 to sgp130 was interfered with specifically by the 4 neutralizing anti-vIL-6 mAbs. Based on the vIL-6 crystal structure, the vIL-6 neutralizing mAbs map outside the binding interface to gp130, suggesting that they either produce allosteric changes or block necessary conformational changes in vIL-6 preceding its binding to gp130. These results document that vIL-6 does not bind IL-6R and suggest that conformational change may be critical to vIL-6 function.
authors.affiliation: Medicine Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA. aokiy@mail.nih.gov
authors.forename: Y
authors.initials: Y
authors.lastname: Aoki
authors.forename: M
authors.initials: M
authors.lastname: Narazaki
authors.forename: T
authors.initials: T
authors.lastname: Kishimoto
authors.forename: G
authors.initials: G
authors.lastname: Tosato
mesh_terms: D000494:Allosteric Regulation; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology; D015703:Antigens, CD / Q000737:chemistry / Q000378:metabolism; D015153:Blotting, Western; D018360:Crystallography, X-Ray; D050822:Cytokine Receptor gp130; D000939:Epitopes / Q000276:immunology; D019288:Herpesvirus 8, Human / Q000235:genetics*; D006801:Humans; D015850:Interleukin-6 / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D009500:Neutralization Tests; D011485:Protein Binding; D011487:Protein Conformation; D019947:Receptors, Interleukin-6 / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D012995:Solubility; D013329:Structure-Activity Relationship; D020349:Surface Plasmon Resonance; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D000939:Epitopes; C494199:IL6ST protein, human; C494200:Il6st protein, mouse; D015850:Interleukin-6; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins; D019947:Receptors, Interleukin-6; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D050822:Cytokine Receptor gp130