Differential binding of human interleukin-1 (IL-1) receptor antagonist to natural and recombinant soluble and cellular IL-1 type I receptors.
A recently described factor, interleukin-1 receptor antagonist binding factor (IL-1raBF), in serum of normal individuals is immunologically related to the interleukin-1 receptor type I (IL-1RI). It is presumably a soluble form of the receptor that binds exclusively to interleukin-1 receptor antagonist (IL-1ra). Recombinant soluble human IL-1RI expressed in COS cells (sIL-1RI) consists of the extracellular part of the receptor and binds all three known IL-1 species but preferentially to Il-1ra. We further characterized the sizes and binding of IL-1raBF and sIL-1RI to IL-1ra by polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate, ligand binding interference analyses, N-glycosidase treatment, concanavalin A affinity chromatography, and with the use of monoclonal antibodies (mAb) to human recombinant IL-1ra. We also evaluated the binding of IL-1ra to cellular IL-1RI on MRC5 fibroblasts and the interference afforded by the soluble receptors. The results show that the protein backbones of IL-1raBF and sIL-1RI are of similar size (approximately 35-40 kDa) and that there are differences in the glycosylation of the two molecules. These carbohydrates were necessary for optimal binding of both molecules to IL-1ra. Both factors blocked binding of IL-1ra to cellular IL-1RI, as did mAb to IL-1ra, but the sites on IL-1ra which bound to the mAb, and to IL-1raBF and sIL-1RI, differed. We conclude that there are important differences between the natural and recombinant forms of soluble IL-1RI and that IL-1ra binds differently to these molecules and to cellular IL-1RI.
Svenson
M
M
Institute for Inflammation Research TA 7544, RHIMA Center, National University Hospital, Copenhagen, Denmark.
Nedergaard
S
S
Heegaard
P M
PM
Whisenand
T D
TD
Arend
W P
WP
Bendtzen
K
K
D000328:Adult; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001667:Binding, Competitive; D002460:Cell Line; D005347:Fibroblasts; D006026:Glycoside Hydrolases / Q000494:pharmacology; D006031:Glycosylation; D006801:Humans; D053590:Interleukin 1 Receptor Antagonist Protein; D008168:Lung; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D017472:Receptors, Interleukin-1 / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D012795:Sialoglycoproteins / Q000378:metabolism*; D012995:Solubility
D000911:Antibodies, Monoclonal; C507763:IL1RN protein, human; D053590:Interleukin 1 Receptor Antagonist Protein; D017472:Receptors, Interleukin-1; D011993:Recombinant Fusion Proteins; D012795:Sialoglycoproteins; D006026:Glycoside Hydrolases