Characterization of a soluble ternary complex formed between human interferon-beta-1a and its receptor chains.
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.
Arduini
R M
RM
Biogen Inc., Cambridge, Massachusetts 02142, USA.
Strauch
K L
KL
Runkel
L A
LA
Carlson
M M
MM
Hronowski
X
X
Foley
S F
SF
Young
C N
CN
Cheng
W
W
Hochman
P S
PS
Baker
D P
DP
D002850:Chromatography, Gel; D019281:Dimerization; D006639:Histidine / Q000737:chemistry; D006801:Humans; D000068556:Interferon beta-1a; D016899:Interferon-beta / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D046911:Macromolecular Substances; D008565:Membrane Proteins; D053633:Receptor, Interferon alpha-beta; D017471:Receptors, Interferon / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D012995:Solubility
C507964:IFNAR1 protein, human; C508033:IFNAR2 protein, human; D046911:Macromolecular Substances; D008565:Membrane Proteins; D017471:Receptors, Interferon; D053633:Receptor, Interferon alpha-beta; D006639:Histidine; D016899:Interferon-beta; D000068556:Interferon beta-1a