Identification of soluble interleukin-1 binding protein in cell-free supernatants. Evidence for soluble interleukin-1 receptor.
This study describes the identification and characterization of a soluble interleukin-1 (IL-1) binding protein in the conditioned media from Raji human B-lymphoma cells. The soluble IL-1 binding material was isolated by IL-1 affinity chromatography, and treatment with trypsin decreased its ability to bind to IL-1 demonstrating its protein nature. The soluble IL-1 binding protein was specific for IL-1 and was able to discriminate between Il-1 alpha and IL-1 beta in a manner analogous to the membrane-bound Raji IL-1 receptor. The specificity of the IL-1 binding protein was further established in two ways. 1) Cell-free supernatants from Raji "receptor-negative" cells did not contain any IL-1 binding protein, thus ruling out nonspecific interactions between IL-1 and a serum or other protein present in the conditioned medium; and 2) the soluble binding protein inhibited IL-1 binding to Raji cells in a dose-dependent manner. Scatchard analysis of IL-1 beta binding showed the dissociation constant (KD) to be 5.1 nM for the soluble IL-1 binding protein compared with 0.8 nM for the membrane-bound IL-1 receptor. Gel chromatography of the soluble binding protein yielded a major peak of IL-1 binding activity with a molecular mass of 35-45 kDa. The characteristics of the soluble IL-1 binding protein described above are consistent with those of the extracellular binding domain of the membrane-bound Raji IL-1 receptor.
Giri
J G
JG
Medical Products Division, E. I. Du Pont De Nemours & Co., Glenolden Laboratory, Pennsylvania 19036.
Newton
R C
RC
Horuk
R
R
D002460:Cell Line; D002462:Cell Membrane / Q000276:immunology; D002999:Clone Cells; D003600:Cytosol / Q000276:immunology; D006801:Humans; D007375:Interleukin-1 / Q000378:metabolism; D007700:Kinetics; D011971:Receptors, Immunologic / Q000302:isolation & purification / Q000378:metabolism*; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins / Q000378:metabolism
D007375:Interleukin-1; D011971:Receptors, Immunologic; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins