title: Efficient purification of recombinant human tumor necrosis factor beta from Escherichia coli yields biologically active protein with a trimeric structure that binds to both tumor necrosis factor receptors.
abstract: A fast and efficient method for medium scale purification of recombinant human tumor necrosis factor beta (rTNF-beta) from Escherichia coli cells is described. The purified rTNF-beta displayed biological activity similar to rTNF-alpha in a WEHI 164 cell cytotoxicity assay. The titration curve of rTNF-beta and elution profiles of rTNF-beta in gel filtration experiments were different from those of rTNF-alpha. However, light scattering and ultra-centrifugation studies showed that both cytokines have trimeric structures in solution at 0.5 mg/ml, with minor differences in the distribution of nontrimeric species. rTNF-beta bound to purified 55- and 75-kDa TNF receptors with high affinity. The binding of rTNF-beta to either receptor was analyzed on Scatchard plots and compared with that of rTNF-alpha.
authors.affiliation: Central Research Units, F. Hoffmann-La Roche Ltd, Basel, Switzerland.
authors.forename: H J
authors.initials: HJ
authors.lastname: Schoenfeld
authors.forename: B
authors.initials: B
authors.lastname: Poeschl
authors.forename: J R
authors.initials: JR
authors.lastname: Frey
authors.forename: H
authors.initials: H
authors.lastname: Loetscher
authors.forename: W
authors.initials: W
authors.lastname: Hunziker
authors.forename: A
authors.initials: A
authors.lastname: Lustig
authors.forename: M
authors.initials: M
authors.lastname: Zulauf
mesh_terms: D001667:Binding, Competitive; D015153:Blotting, Western; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D008233:Lymphotoxin-alpha / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D018124:Receptors, Tumor Necrosis Factor; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D008233:Lymphotoxin-alpha; D011956:Receptors, Cell Surface; D018124:Receptors, Tumor Necrosis Factor; D011994:Recombinant Proteins