title: Purification of receptor complexes of interleukin-10 stoichiometry and the importance of deglycosylation in their crystallization.
abstract: Interleukin-10 (IL-10) is a pleiotropic immunosuppressive cytokine that has a wide range of effects in controlling inflammatory responses. Viral IL-10 (vIL-10) is a homologue of human IL-10 (hIL-10) produced by Epstein-Barr virus (EBV). Both hIL-10 and vIL-10 bind to the soluble extracellular fragment of the cytokine receptor IL-10R1 (shIL-10R1). The stoichiometry of the vIL-10 : shIL-10R1 complex has been found to be the same as hIL-10 : shIL-10R1, with two vIL-10 dimers binding to four shIL-10R1 monomers. Complexes of both hIL-10 and vIL-10 with glycosylated shIL-10R1 could not be crystallized. Controlled deglycosylation using peptide : N-glycosidase F and endo-beta-N-acetylglucosaminidase F3 resulted in the formation of crystals of both hIL-10 : shIL-10R1 and vIL-10 : shIL-10R1 complexes, indicating that the difficulty in the crystal formation was largely due to the presence of complex carbohydrate side chains. The availability of the structure of the ligand-receptor complexes should facilitate our understanding of the basis of the interaction between IL-10 and the IL-10 receptor.
authors.affiliation: Macromolecular Structure Laboratory, NCI-Frederick Cancer Research and Development Center, ABL-Basic Research Program, Frederick, MD, USa.
authors.forename: D M
authors.initials: DM
authors.lastname: Hoover
authors.forename: C
authors.initials: C
authors.lastname: Schalk-Hihi
authors.forename: C C
authors.initials: CC
authors.lastname: Chou
authors.forename: S
authors.initials: S
authors.lastname: Menon
authors.forename: A
authors.initials: A
authors.lastname: Wlodawer
authors.forename: A
authors.initials: A
authors.lastname: Zdanov
mesh_terms: D002850:Chromatography, Gel; D003460:Crystallization; D018360:Crystallography, X-Ray; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation; D006801:Humans; D016753:Interleukin-10 / Q000737:chemistry* / Q000302:isolation & purification; D018123:Receptors, Interleukin / Q000737:chemistry* / Q000302:isolation & purification; D053701:Receptors, Interleukin-10; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D018123:Receptors, Interleukin; D053701:Receptors, Interleukin-10; D011994:Recombinant Proteins; D016753:Interleukin-10