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Characterization of a soluble ternary complex formed between human interferon-β-1a and its receptor chains

Published online by Cambridge University Press:  01 September 1999

ROBERT M. ARDUINI
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
KATHRYN L. STRAUCH
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
LAURA A. RUNKEL
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
MELISSA M. CARLSON
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
XIAOPING HRONOWSKI
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
SUSAN F. FOLEY
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
CARMEN N. YOUNG
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
WENJIE CHENG
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
PAULA S. HOCHMAN
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
DARREN P. BAKER
Affiliation:
Biogen Inc., 14 Cambridge Center, Cambridge, Massachusetts 02142
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Abstract

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-β-1a (IFN-β-1a) were studied by gel filtration and by cross-linking. By gel filtration, no stable binary complexes between IFN-β-1a and IFNAR1, or between IFNAR1 and IFNAR2 were detected. However, a stable binary complex formed between IFN-β-1a and IFNAR2. Analysis of binary complex formation using various molar excesses of IFN-β-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 M>r consistent with the sum of its two individual components. Gel filtration of a mixture of IFNAR1 and the IFN-β-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-β-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-β-1a with IFNAR2, followed by the association of IFNAR1 with the preformed binary complex. The ability to produce the IFN-β-1a/IFNAR2 and IFN-β-1a/IFNAR1/IFNAR2 complexes make them attractive candidates for X-ray crystallography studies aimed at determining the molecular interactions between IFN-β-1a and its receptor.

Type
Research Article
Copyright
© 1999 The Protein Society

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