One interferon gamma receptor binds one interferon gamma dimer.
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.
Fountoulakis
M
M
Hoffman-La Roche Limited, Central Research Units, Basel, Switzerland.
Juranville
J F
JF
Maris
A
A
Ozmen
L
L
Garotta
G
G
D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007371:Interferon-gamma / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D010084:Oxidation-Reduction; D011487:Protein Conformation; D011971:Receptors, Immunologic / Q000737:chemistry / Q000378:metabolism*; D017471:Receptors, Interferon; D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D013861:Thiocyanates / Q000494:pharmacology
D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D011971:Receptors, Immunologic; D017471:Receptors, Interferon; D011994:Recombinant Proteins; D013861:Thiocyanates; D012967:Sodium Dodecyl Sulfate; C024553:sodium thiocyanate; D007371:Interferon-gamma