Enrichment and initial characterization of the solubilized receptor for mouse gamma interferon.
The work reported here constitutes a first step in characterizing the receptor for mouse gamma interferon at the biochemical level. The myelomonocytic cell line, WEHI-3, was the source of starting material. Iodinated recombinant mouse gamma interferon incubated with WEHI-3 cells, as well as membranes prepared from them, bound specifically to a single class of sites with a Kd of 7 x 10(-9)M. Membranes were solubilized with the non-ionic detergent octyl-beta-D-glucopyranoside. As solubilization proceeded, binding activity could be assayed by precipitating the receptor with acetone in the presence of egg phosphatidylcholine liposomes. The Kd of the receptor in association with liposomes was 13 nM. Again here, only a single class of binding activity was found, and specificity for gamma, compared to other interferons, was maintained. This is the first time that the receptor for mouse gamma interferon has been solubilized and recovered in functional form. Further characterization included at least a 200-fold enrichment of binding activity by ligand affinity chromatography, resulting in the identification of a 95 kDa protein as the most likely candidate for either the receptor or a binding subunit thereof.
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D011232:Chemical Precipitation; D002846:Chromatography, Affinity; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D005960:Glucosides; D006098:Granulocytes / Q000378:metabolism; D007371:Interferon-gamma / Q000378:metabolism*; D008081:Liposomes / Q000378:metabolism; D051379:Mice; D008970:Molecular Weight; D009000:Monocytes / Q000378:metabolism; D011971:Receptors, Immunologic / Q000378:metabolism*; D017471:Receptors, Interferon; D011994:Recombinant Proteins / Q000378:metabolism; D012995:Solubility
D003902:Detergents; D005960:Glucosides; D008081:Liposomes; D011971:Receptors, Immunologic; D017471:Receptors, Interferon; D011994:Recombinant Proteins; C018619:octyl-beta-D-glucoside; D007371:Interferon-gamma