title: Purification and sequencing of glycosylation variants of BSF-1, as a MAF, from the EL-4 leukaemia cell line.
abstract: Macrophage activation activity was characterized from a PMA-induced subclone of the murine EL-4 leukaemic cell line. The MAF was purified from the cell line culture supernatant by concentration, CM-Sepharose and lentil lectin Sepharose chromatography, AcA 54 gel filtration, Mono Q FPLC and reverse phase HPLC. Four protein bands of different abundance were observed on SDS-PAGE with molecular weights of 17,500 to 21,000 Da. Three of the four proteins were sequenced from the N-terminal and shared homology with the published sequence of BSF-1. Variation of the molecular weight due to glycosylation was demonstrated by N-glycanase treatment, all four proteins gave a band of 14,200 Da after deglycosylation. Both glycosylated and deglycosylated forms of BSF-1 were equally active in the MAF assay. A monoclonal antibody to BSF-1 neutralized 80% of the activity from crude culture supernatants in the MAF assay. These studies have indicated that BSF-1 is the major, if not the only, MAF activity from this particular subline of the murine EL-4 leukaemic cell line.
authors.affiliation: Celltech Limited, Berkshire, UK.
authors.forename: C
authors.initials: C
authors.lastname: Sutton
authors.forename: P
authors.initials: P
authors.lastname: Depledge
authors.forename: L
authors.initials: L
authors.lastname: Bawden
authors.forename: A
authors.initials: A
authors.lastname: Carne
authors.forename: M
authors.initials: M
authors.lastname: Meltzer
authors.forename: V
authors.initials: V
authors.lastname: Newton
authors.forename: L
authors.initials: L
authors.lastname: Vodinelich
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D006031:Glycosylation; D015847:Interleukin-4; D007378:Interleukins / Q000302:isolation & purification*; D004915:Leukemia, Erythroblastic, Acute / Q000276:immunology*; D008222:Lymphokines / Q000302:isolation & purification*; D016215:Macrophage-Activating Factors; D008969:Molecular Sequence Data; D008970:Molecular Weight; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000276:immunology
chemical_list: D007378:Interleukins; D008222:Lymphokines; D016215:Macrophage-Activating Factors; D015847:Interleukin-4; D013755:Tetradecanoylphorbol Acetate