Stoichiometry of interaction between interferon gamma and its receptor.
The biological response of interferon gamma is mediated by binding to a specific cell-surface receptor. We investigated the stoichiometry of this binding using soluble receptors produced in prokaryotic and eukaryotic expression systems comprising the extracellular ligand-binding domain of the native protein. The ligand-receptor complexes were analyzed by cross-linking, chromatography, analytical ultracentrifugation and laser-light scattering. Cross-linking and chromatography showed that the stoichiometry of the interaction between ligand and receptor depends on the molar ratios of the two components mixed. All approaches confirmed that mixtures of ligand-receptor complexes are formed with one interferon-gamma dimer bound by one or two receptors. The soluble receptor produced in Escherichia coli mainly showed a ligand/receptor stoichiometry of 1:1, while the receptors produced in eukaryotic cells showed a stoichiometry of binding of 1:2. This apparent discrepancy is most likely due to the conformational heterogeneity of the Escherichia-coli-derived protein.
Fountoulakis
M
M
F. Hoffmann-La Roche Ltd., Pharmaceutical Research-New Technologies, Basel, Switzerland.
Zulauf
M
M
Lustig
A
A
Garotta
G
G
D000595:Amino Acid Sequence; D003432:Cross-Linking Reagents; D006031:Glycosylation; D066298:In Vitro Techniques; D007371:Interferon-gamma / Q000378:metabolism*; D008024:Ligands; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D012542:Scattering, Radiation; D012995:Solubility; D013329:Structure-Activity Relationship; D014461:Ultracentrifugation
D003432:Cross-Linking Reagents; D008024:Ligands; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D007371:Interferon-gamma