One interferon gamma receptor binds one interferon gamma dimer

• Published in: The Journal of biological chemistry Vol. 265; no. 32; pp. 19758 - 19767
• Main Authors: Fountoulakis, M, Juranville, J F, Maris, A, Ozmen, L, Garotta, G
• Format: Journal Article
• Language: English
• Published: Bethesda, MD Elsevier Inc 15.11.1990; American Society for Biochemistry and Molecular Biology
• Subjects: Biological and medical sciences; Cell Line; Cell Membrane - metabolism; Cell receptors; Cell structures and functions; Chromatography, Gel; Cross-Linking Reagents; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Humans; Interferon-gamma - chemistry; Interferon-gamma - metabolism; Kinetics; Macromolecular Substances; Miscellaneous; Molecular and cellular biology; Molecular Weight; Oxidation-Reduction; Protein Conformation; Receptors, Immunologic - chemistry; Receptors, Immunologic - metabolism; Receptors, Interferon; Recombinant Proteins; Sodium Dodecyl Sulfate - pharmacology; Thiocyanates - pharmacology; Stoichiometry; Binding capacity; Cytokine; Gel filtration; Molecular interaction; Crosslinking; Dimer; Interferon; Kinetics; Inhibition; Molecular dissociation; Biological receptor
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(17)45437-8

We investigated the stoichiometry of the interferon gamma and interferon gamma receptor interaction, using recombinant interferon gamma and recombinant soluble interferon gamma receptor, applying chemical cross-linking and chromatographic techniques, and analyzing the resulting products in denaturing polyacrylamide gels. Interferon gamma cross-linked to itself produced a major band of an apparent molecular mass of 34 kDa, which suggests that it exists as a dimer in physiological buffer and which agrees with published data. Soluble interferon gamma receptor cross-linked to itself produced mainly a 28-kDa band, suggesting that the interferon gamma receptor exists as a monomer. Interferon gamma cross-linked to the soluble interferon gamma receptor resulted in the formation of two main products of apparent molecular masses of 60 and 44 kDa. The predominant 60-kDa band resulted from the cross-linking of one interferon gamma dimer (34 kDa) to one interferon gamma receptor molecule (27 kDa). The 44-kDa band was formed by the cross-linking of one interferon gamma molecule to one interferon gamma receptor. Kinetic studies showed that the cross-linking of interferon gamma dimer to the soluble receptor proceeds through the intermediate formed by cross-linking one molecule of the interferon gamma dimer to the receptor. Reducing and dissociating agents inhibited complex formation. When chromatographed on Sephadex G-100, interferon gamma was eluted as a protein of 34-kDa molecular mass, the soluble interferon gamma receptor as a protein of 40 kDa, and their mixture was eluted in one peak corresponding to an apparent molecular mass of 73 kDa. Sodium dodecyl sulfate-polyacrylamide gel analysis of the eluted mixture showed the presence of both interferon gamma and interferon gamma receptor at a ratio of 2:1. The found results suggest that the interferon gamma receptor binds interferon gamma as a dimer.