Murine interleukin 1 receptor. Direct identification by ligand blotting and purification to homogeneity of an interleukin 1-binding glycoprotein.
Functional receptors (IL1-R) for the proinflammatory cytokine interleukin 1 (IL1) were solubilized from plasma membranes of the NOB-1 subclone of murine EL4 6.1 thymoma cells using the zwitterionic detergent 3[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS). Membrane extracts were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose membranes, and "ligand blotted" with 125I-labeled recombinant human IL1 alpha in order to reveal proteins capable of specifically binding IL1. A single polydisperse polypeptide of Mr approximately equal to 80,000 was identified in this way, which bound IL1 alpha and IL1 beta with the same affinity as the IL1-R on intact NOB-1 cells (approximately equal to 10(-10) M). The IL1-binding polypeptide was only seen in membranes from IL1-R-bearing cells and did not react with interleukin 2, tumor necrosis factor alpha, or interferon. IL1-R was purified to apparent homogeneity from solubilized NOB-1 membranes by affinity chromatography on wheat germ agglutinin-Sepharose and IL1 alpha-Sepharose. Gel electrophoresis and silver staining of purified preparations revealed a single protein of Mr approximately equal to 80,000 which reacted positively in the ligand-blotting procedure and which we identify as the ligand-binding moiety of the murine IL1-R. Purified IL1-R exhibited the same affinity and specificity as the receptor on intact cells. The relationship of this protein to proteins identified by covalent cross-linking studies is discussed.
Bird
T A
TA
Cytokine Biochemistry Group, Strangeways Research Laboratory, Cambridge, United Kingdom.
Gearing
A J
AJ
Saklatvala
J
J
D000818:Animals; D001667:Binding, Competitive; D002462:Cell Membrane / Q000032:analysis; D002793:Cholic Acids; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007375:Interleukin-1 / Q000378:metabolism; D007457:Iodine Radioisotopes; D051379:Mice; D008970:Molecular Weight; D011948:Receptors, Antigen, T-Cell; D011971:Receptors, Immunologic / Q000032:analysis* / Q000378:metabolism; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins / Q000378:metabolism; D012995:Solubility; D013945:Thymoma / Q000032:analysis*; D013953:Thymus Neoplasms / Q000032:analysis*; D014407:Tumor Cells, Cultured
D002793:Cholic Acids; D007375:Interleukin-1; D007457:Iodine Radioisotopes; D011948:Receptors, Antigen, T-Cell; D011971:Receptors, Immunologic; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate