title: Identification and characterization of a C-terminally extended form of recombinant murine IL-6.
abstract: Murine interleukin-6 (mIL-6) was expressed in Escherichia coli in the insoluble fraction of cell lysates. Approximately equal amounts of two polypeptide species, reactive with anti-IL-6 antibodies, were produced. The two forms of mIL-6 were isolated and found to have identical N-terminal sequences initiated by Met-Phe-Pro-Thr-Ser-Gln-. Peptide mapping after endoproteinase glu-C digestion led to isolation and characterization of the C-terminal peptides from each of the two forms and allowed the source of the heterogeneity to be identified as a C-terminal addition of three amino acids, Gln-Lys-Leu, to authentic mIL-6. Inspection of the nucleotide sequence of the plasmid containing the mIL-6 gene and expression of the plasmid in other strains suggested that the addition of three amino acids was caused by a readthrough of the termination codon arising from an unexpected suppressor mutation in the original host strain. Although the C-terminus of IL-6 is critical for the activity of this cytokine, the IL-6 variant with extended C-terminus was fully active in two separate bioassays. This suggests that the additional amino acids do not disrupt the structure or function of this important region of the molecule.
authors.affiliation: Department of Molecular Genetics, Pfizer Inc., Groton, CT 06340.
authors.forename: D E
authors.initials: DE
authors.lastname: Danley
authors.forename: C A
authors.initials: CA
authors.lastname: Strick
authors.forename: L C
authors.initials: LC
authors.lastname: James
authors.forename: A J
authors.initials: AJ
authors.lastname: Lanzetti
authors.forename: I G
authors.initials: IG
authors.lastname: Otterness
authors.forename: H E
authors.initials: HE
authors.lastname: Grenett
authors.forename: G M
authors.initials: GM
authors.lastname: Fuller
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D015850:Interleukin-6 / Q000737:chemistry*; D051379:Mice; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D011994:Recombinant Proteins / Q000737:chemistry; D016415:Sequence Alignment
chemical_list: D015850:Interleukin-6; D011994:Recombinant Proteins