Glycoprotein 130 (gp130) is a type I transmembrane protein and serves as the common signal-transducing receptor subunit of the interleukin-6-type cytokines. Whereas the membrane-distal half of the gp130 extracellular part confers ligand binding and has been subject to intense investigation, the structural and functional features of its membrane-proximal half are poorly understood. On the basis of predictions of tertiary structure, the membrane-proximal part consists of three fibronectin-type-III-like domains D4, D5 and D6. Here we describe the bacterial expression of the polypeptides predicted to comprise each of these three domains. The recombinant proteins were refolded from solubilized inclusion bodies in vitro, purified to homogeneity and characterized by means of size-exclusion chromatography and CD spectroscopy. For the first time the prediction of three individual membrane-proximal protein domains for gp130has been verified experimentally. The three domains do not show intermediate-affinity or high-affinity interactions between each other. Mapping of a neutralizing gp130 monoclonal antibody against D4 suggested a particular functional role of this domain for gp130 activation, because above that an intrinsic tendency for low-affinity oligomerization was demonstrated for D4.
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June 2001
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Research Article|
May 24 2001
Signal transducer gp130: biochemical characterization of the three membrane-proximal extracellular domains and evaluation of their oligomerization potential
Stefan PFLANZ;
Stefan PFLANZ
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Thomas KERNEBECK;
Thomas KERNEBECK
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Bernd GIESE;
Bernd GIESE
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Andreas HERRMANN;
Andreas HERRMANN
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Michael PACHTA-NICK;
Michael PACHTA-NICK
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Jürgen STAHL;
Jürgen STAHL
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Axel WOLLMER;
Axel WOLLMER
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Peter C. HEINRICH;
Peter C. HEINRICH
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Gerhard MÜLLER-NEWEN;
Gerhard MÜLLER-NEWEN
1
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
1To whom correspondence should be addressed (e-mail mueller-newen@rwth-aachen.de).
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Joachim GRÖTZINGER
Joachim GRÖTZINGER
Department of Biochemistry, RWTH Aachen, Pauwelsstrasse 30, 52074 Aachen, Germany
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Publisher: Portland Press Ltd
Received:
January 02 2001
Revision Received:
February 28 2001
Accepted:
March 27 2001
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2001
2001
Biochem J (2001) 356 (2): 605–612.
Article history
Received:
January 02 2001
Revision Received:
February 28 2001
Accepted:
March 27 2001
Citation
Stefan PFLANZ, Thomas KERNEBECK, Bernd GIESE, Andreas HERRMANN, Michael PACHTA-NICK, Jürgen STAHL, Axel WOLLMER, Peter C. HEINRICH, Gerhard MÜLLER-NEWEN, Joachim GRÖTZINGER; Signal transducer gp130: biochemical characterization of the three membrane-proximal extracellular domains and evaluation of their oligomerization potential. Biochem J 1 June 2001; 356 (2): 605–612. doi: https://doi.org/10.1042/bj3560605
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