Radiation inactivation of human gamma-interferon: cellular activation requires two dimers.
gamma-Interferon (IFN-gamma) is a 17-kDa broad-spectrum cytokine which exerts its effects on a variety of target cells through its interaction with the IFN-gamma receptor. Although physicochemical studies of Escherichia coli-derived IFN-gamma, as well as its crystal structure, demonstrate that it is a homodimer in solution (M(r) 34,000), previous radiation inactivation studies yielded a functional size for IFN-gamma of 63-73 kDa in an antiviral assay. To understand the relationship between the solution form of IFN-gamma and the moiety that actually binds to the cellular receptor and activates cells, we examined irradiated nonradioactive and 32P-labeled IFN-gamma for its migration in SDS/polyacrylamide gels (to determine its physical integrity), its binding to cells, its reactivity in an ELISA, and its antiviral activity. The functional size of IFN-gamma differed in the assays, being 22 +/- 2 kDa for the physical destruction of IFN-gamma, 56 +/- 2 kDa for the cellular binding assay, 45-50 kDa for reactivity in the ELISA, and 72 +/- 6 kDa for antiviral activity. The results from the binding assays constitute direct evidence that IFN-gamma binds to its cellular receptor as a dimer. However, for antiviral activity, the functional mass is equivalent to a tetramer. This is consistent with models involving ligand-induced receptor dimerization, whereby two dimers acting in concert (equivalent to the target size of a tetramer) are required to activate cells in the antiviral assay.
Department of Molecular Genetics and Microbiology, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway 08854.
J A
JA
Langer
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Rashidbaigi
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G
Garotta
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E
Kempner
D000255:Adenosine Triphosphate / Q000378:metabolism; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D004686:Encephalomyocarditis virus / Q000187:drug effects*; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli; D006801:Humans; D007371:Interferon-gamma / Q000737:chemistry / Q000528:radiation effects* / Q000633:toxicity*; D015473:Leukemia, Promyelocytic, Acute; D046911:Macromolecular Substances; D010761:Phosphorus Radioisotopes; D017471:Receptors, Interferon / Q000378:metabolism; D011994:Recombinant Proteins; D014407:Tumor Cells, Cultured; D014721:Vesicular stomatitis Indiana virus / Q000187:drug effects*; D000096743:Interferon gamma Receptor
D000255:Adenosine Triphosphate; D007371:Interferon-gamma; D046911:Macromolecular Substances; D010761:Phosphorus Radioisotopes; D017471:Receptors, Interferon; D011994:Recombinant Proteins; D000096743:Interferon gamma Receptor