Characterization of a soluble ternary complex formed between human interferon-beta-1a and its receptor chains
| Autor: | Melissa M. Carlson, Robert M. Arduini, Kathryn L. Strauch, Xiaoping Hronowski, Susan F. Foley, Paula S. Hochman, Laura Runkel, Darren P. Baker, Wenjie Cheng, Carmen N. Young |
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| Rok vydání: | 1999 |
| Předmět: |
Stereochemistry
Macromolecular Substances Size-exclusion chromatography Receptor Interferon alpha-beta Biochemistry medicine Humans Histidine Solubility Molecular Biology Ternary complex Receptors Interferon Chemistry Interferon beta-1a Membrane Proteins Interferon-beta Membrane protein Reagent Chromatography Gel Dimerization Stoichiometry medicine.drug Research Article |
| Zdroj: | Protein science : a publication of the Protein Society. 8(9) |
| ISSN: | 0961-8368 |
| Popis: | The extracellular portions of the chains that comprise the human type I interferon receptor, IFNAR1 and IFNAR2, have been expressed and purified as recombinant soluble His-tagged proteins, and their interactions with each other and with human interferon-beta-1a (IFN-beta-1a) were studied by gel filtration and by cross-linking. By gel filtration, no stable binary complexes between IFN-beta-1a and IFNAR1, or between IFNAR1 and IFNAR2 were detected. However, a stable binary complex formed between IFN-beta-1a and IFNAR2. Analysis of binary complex formation using various molar excesses of IFN-beta-1a and IFNAR2 indicated that the complex had a 1:1 stoichiometry, and reducing SDS-PAGE of the binary complex treated with the cross-linking reagent dissucinimidyl glutarate (DSG) indicated that the major cross-linked species had an apparent Mr consistent with the sum of its two individual components. Gel filtration of a mixture of IFNAR1 and the IFN-beta-1a/IFNAR2 complex indicated that the three proteins formed a stable ternary complex. Analysis of ternary complex formation using various molar excesses of IFNAR1 and the IFN-beta-1a/IFNAR2 complex indicated that the ternary complex had a 1:1:1 stoichiometry, and reducing SDS-PAGE of the ternary complex treated with DSG indicated that the major cross-linked species had an apparent Mr consistent with the sum of its three individual components. We conclude that the ternary complex forms by the sequential association of IFN-beta-1a with IFNAR2, followed by the association of IFNAR1 with the preformed binary complex. The ability to produce the IFN-beta-1a/IFNAR2 and IFN-beta-1a/IFNAR1/IFNAR2 complexes make them attractive candidates for X-ray crystallography studies aimed at determining the molecular interactions between IFN-beta-1a and its receptor. |
| Databáze: | OpenAIRE |
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