Affinity purification and chemical analysis of the interleukin-1 receptor

J Biol Chem. 1988 Feb 25;263(6):2870-7.

Abstract

Interleukins-1 alpha and -1 beta regulate the metabolism of cells through a common plasma membrane receptor protein. In this study, it is demonstrated that the interleukin-1 (IL-1) receptor from detergent solutions of EL-4 cells can be stably adsorbed to nitrocellulose with full retention of IL-1 binding activity. This assay system was used to monitor the purification of the IL-1 receptor and to investigate the effects of several chemical modifications on receptor binding activity. IL-1 receptors extracted from EL-4 6.1 C10 cells can be bound to and specifically eluted from IL-1 alpha coupled to Sepharose. The affinity chromatography method resulted in the identification by polyacrylamide gel electrophoresis and silver staining of a protein of Mr 82,000 that was present in fractions exhibiting IL-1 binding activity. Experiments in which the cell-surface proteins of EL-4 cells were radiolabeled and 125I-labeled receptor was purified by affinity chromatography suggested that the Mr 82,000 protein was expressed on the plasma membrane. N-Glycanase treatment of this material showed that 23-35% of the total Mr (82,000) of the receptor is N-linked carbohydrate.

MeSH terms

  • Chromatography, Affinity
  • Detergents
  • Glycoside Hydrolases / metabolism
  • Humans
  • Hydrogen-Ion Concentration
  • Interleukin-1 / metabolism
  • Kinetics
  • Methods
  • Receptors, Immunologic / analysis
  • Receptors, Immunologic / isolation & purification*
  • Receptors, Interleukin-1
  • Tumor Cells, Cultured / analysis

Substances

  • Detergents
  • Interleukin-1
  • Receptors, Immunologic
  • Receptors, Interleukin-1
  • Glycoside Hydrolases
  • glycanase