title: Role of disulfide bonds in biologic activity of human interleukin-6.
abstract: We have examined the functional importance of the two disulfide bonds formed by the four conserved cysteines of human interleukin (IL-6). Using a bacterial expression system, we have synthesized a series of recombinant IL-6 mutants in which the constituent cysteines of the first (Cys45-Cys51), second (Cys74-Cys84), or both disulfide bonds of recombinant human interleukin-6 were replaced by other amino acids. Each mutant was partially purified and tested in four representative bioassays. While mutants lacking Cys45 and Cys51 retained activity similar to nonmutated recombinant IL-6, the activity of mutants lacking Cys74 and Cys84 was significantly reduced, especially in assays involving human cell lines. These results indicate that the first disulfide bond of human interleukin-6 is not required for maintenance of normal biologic activity. However, the fact that mutants lacking Cys45 and Cys51 were more active than corresponding cysteine-free mutants indicates that the disulfide bond formed by these residues contributes to biologic activity in the absence of the second disulfide bond. Competition binding studies with representative mutants indicate that their affinity for the human IL-6 receptor parallels their biologic activities on human cells.
authors.affiliation: Department of Pathology, University of North Carolina, Chapel Hill 27599-7525.
authors.forename: J N
authors.initials: JN
authors.lastname: Snouwaert
authors.forename: F W
authors.initials: FW
authors.lastname: Leebeek
authors.forename: D M
authors.initials: DM
authors.lastname: Fowlkes
mesh_terms: D001667:Binding, Competitive; D002460:Cell Line; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry*; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D015850:Interleukin-6 / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D016296:Mutagenesis; D011994:Recombinant Proteins
chemical_list: D004220:Disulfides; D015850:Interleukin-6; D011994:Recombinant Proteins; D003545:Cysteine