title: High-level expression of human BSF-2/IL-6 cDNA in Escherichia coli using a new type of expression-preparation system.
abstract: BSF-2 (B cell stimulatory factor-2/IL-6) is a member of the lymphokine family and responsible for B cell differentiation. Expression plasmids of human BSF-2 cDNA were constructed using a trp promotor/operator and a trpA terminator. In an extract of Escherichia coli HB101 holding "direct" expression plasmid pBSF-2D, activity of BSF-2 was detected, but overproduction was not observed. A "fused" expression system was therefore developed to prepare the recombinant protein. In this system, cDNA was expressed as a fused protein with human IL-2 N-terminal peptide. In the case of the fused BSF-2 expression plasmid, pBSF-2F, inclusion bodies were observed and overproduction of the protein occurred. As this fused protein had a Phe-Arg-Ala sequence at the junction of hIL-2 and BSF-2, it was possible to process mature BSF-2 from the fused BSF-2 by treatment with kallikrein and aminopeptidase P. From 1 liter of E. coli culture, 45 mg of mature BSF-2 was purified; it had a relative biological activity equal to that of natural BSF-2 purified from T cells.
authors.affiliation: Central Research Laboratories, Ajinomoto Co., Inc., Kanagawa.
authors.forename: N
authors.initials: N
authors.lastname: Tonouchi
authors.forename: N
authors.initials: N
authors.lastname: Oouchi
authors.forename: N
authors.initials: N
authors.lastname: Kashima
authors.forename: M
authors.initials: M
authors.lastname: Kawai
authors.forename: K
authors.initials: K
authors.lastname: Nagase
authors.forename: A
authors.initials: A
authors.lastname: Okano
authors.forename: H
authors.initials: H
authors.lastname: Matsui
authors.forename: K
authors.initials: K
authors.lastname: Yamada
authors.forename: T
authors.initials: T
authors.lastname: Hirano
authors.forename: T
authors.initials: T
authors.lastname: Kishimoto
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004261:DNA Replication; D004262:DNA Restriction Enzymes; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005822:Genetic Vectors; D006801:Humans; D015850:Interleukin-6; D007378:Interleukins / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids
chemical_list: D015850:Interleukin-6; D007378:Interleukins; D004262:DNA Restriction Enzymes