title: Modulation of the biologic activities of IgE-binding factor. IV. Identification of glycosylation-enhancing factor as a kallikrein-like enzyme.
abstract: Stimulation of normal rat splenic T cells with pertussigen (lymphocytosis-promoting factor, LPF, from Bordetella pertussis) resulted in the release of a soluble factor that enhanced the glycosylation of IgE-binding factors during their biosynthesis. The soluble factor was detected by the ability of a culture filtrate of LPF-stimulated spleen cells to switch a T cell hybridoma, 23A4, from the formation of unglycosylated IgE-binding factor to the formation of glycosylated IgE-binding factor. The glycosylation-enhancing factor (GEF) had affinity for D-galactose, and the binding of the factor to hybridoma cells via a cell surface galactose was essential for modulation of IgE-binding factors. The GEF was inactivated by irreversible inhibitors of serine proteases such as phenylmethylsulfonyl fluoride, diisopropylfluorophosphate, and p-nitrophenyl ethylpentylphosphonate but was not affected by nonphosphorylating analogues of the organophosphorus compounds. Benzamidine, a competitive and reversible inhibitor of trypsin, also inhibited the glycosylation of IgE-binding factors by GEF. The factor could be purified by absorption to p-aminobenzamidine agarose followed by elution with benzamidine. The capacity of GEF to enhance the glycosylation of IgE-binding factors was inhibited by synthetic substrates of trypsin but not by substrates of chymotrypsin, indicating that GEF is a trypsin-like enzyme. Indeed, trypsin, plasmin, and kallikrein enhanced the glycosylation of IgE-binding factors during their biosynthesis. An inhibitor of trypsin-like enzyme(s), N-alpha-p-tosyl-L-lysine chloromethylketone (TLCK), inhibited trypsin and plasmin but not kallikrein, and TLCK failed to inhibit the GEF-mediated enhancement of glycosylation. It was also found that bradykinin, the biologically active product of cleavage of kininogen by kallikrein, enhanced the glycosylation of IgE-binding factors. The results indicate that GEF is a kallikrein-like enzyme.
mesh_terms: D000818:Animals; D001550:Benzamidines / Q000494:pharmacology; D001667:Binding, Competitive; D001798:Blood Proteins / Q000032:analysis / Q000302:isolation & purification / Q000502:physiology*; D004187:Disaccharides / Q000494:pharmacology; D007610:Kallikreins; D008198:Lymph Nodes / Q000166:cytology; D008222:Lymphokines / Q000096:biosynthesis / Q000378:metabolism* / Q000502:physiology; D009005:Monosaccharides / Q000494:pharmacology; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D029584:Prostatic Secretory Proteins; D011480:Protease Inhibitors / Q000494:pharmacology; D051381:Rats; D014357:Trypsin / Q000494:pharmacology
chemical_list: D001550:Benzamidines; D001798:Blood Proteins; D004187:Disaccharides; D008222:Lymphokines; D009005:Monosaccharides; D029584:Prostatic Secretory Proteins; D011480:Protease Inhibitors; C043076:beta-microseminoprotein; C011850:immunoenhancing factor; C013059:immunoglobulin-binding factors; D010664:Phenylmethylsulfonyl Fluoride; D007610:Kallikreins; D014357:Trypsin; C032157:benzamidine