title: The signal transducer gp130--bacterial expression, refolding and properties of the carboxy-terminal domain of the cytokine-binding module.
abstract: Gp130 is the signal transducing receptor subunit of the so-called interleukin-6-type cytokines. This transmembrane protein is a member of the cytokine-receptor superfamily predicted to consist of six fibronectin-type-III-like domains in its extracellular part. The second and the third domain constitute the so-called cytokine-binding module. Domain 2 is characterized by a set of four conserved Cys residues, domain 3 by a conserved WSXWS motif. As a first approach to a more detailed characterization of the cytokine-binding domains of human gp130, we have expressed in Escherichia coli two forms of domain 3 differing in length. Both proteins were purified and refolded in a single step applying size-exclusion chromatography. According to the rotational correlation times deduced from fluorescence anisotropy decay, they do not form aggregates. CD and fluorescence spectroscopy were used to study thermal unfolding and denaturation by guanidinium hydrochloride. It was shown that N- and C-terminal extension by residues of the adjacent hinge regions substantially increase the thermal stability of the domain, which is conceivable from a molecular model. These results are the basis for further structural investigation by NMR spectroscopy.
authors.affiliation: Institut für Biochemie, Rheinisch-Westfälische Technische Hochschule Aachen, Germany.
authors.forename: G
authors.initials: G
authors.lastname: Müller-Newen
authors.forename: S
authors.initials: S
authors.lastname: Pflanz
authors.forename: U
authors.initials: U
authors.lastname: Hassiepen
authors.forename: J
authors.initials: J
authors.lastname: Stahl
authors.forename: A
authors.initials: A
authors.lastname: Wollmer
authors.forename: P C
authors.initials: PC
authors.lastname: Heinrich
authors.forename: J
authors.initials: J
authors.lastname: Grötzinger
mesh_terms: D001665:Binding Sites; D002942:Circular Dichroism; D016207:Cytokines / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D015850:Interleukin-6 / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D017510:Protein Folding; D018121:Receptors, Cytokine / Q000737:chemistry*; D015398:Signal Transduction; D013050:Spectrometry, Fluorescence
chemical_list: D016207:Cytokines; D015850:Interleukin-6; D018121:Receptors, Cytokine