Immobilization of galactosyltransferase and continuous galactosylation of glycoproteins in a reactor |
| |
Authors: | Robert Schneider Mario Hammel Eric G Berger Oreste Ghisalba Jakob Nueesch Daniel Gygax |
| |
Affiliation: | (1) Central Research Laboratories, Ciba-Geigy AG, Klybeckstr. 141, 4057 Basel, Switzerland;(2) Institute of Physiology, University of Zurich, Winterthurerstr. 190, 8057 Zurich, Switzerland;(3) Pharma Division, Ciba-Geigy AG, Klybeckstr. 141, 4057 Basel, Switzerland |
| |
Abstract: | We immobilized human milk galactosyltransferase covalently to CNBr- and tresylchloride-activated Sepharose. The enzyme was also immobilized non-covalently to Concanavalin A-Sepharose and to monoclonal anti-galactosyltransferase antibodies which were boundvia their Fc-fragment to Protein G-Sepharose. With the covalent methods, up to 72% of the enzyme could be bound to the carrier, but more than 90% of the specific activity was lost. In contrast, non-covalent immobilization yielded only about 50% immobilization efficiency, but 21% and 25% of specific activity, respectively, could be recovered. The stability of immobilized galactosyltransferase as compared to native enzyme was considerably increased: at room temperature, 55% of initial immobilized activity was lost after 65 hours compared to 95% of loss of soluble enzyme activity. Immobilized galactosyltransferase was then used for continuous galactosylation of the glycoproteins ovalbumin, endo H-treated yeast invertase and bovine serum albumin-N-acetylglucosamine in a slurry reactor. 55%, 35% and 25%, respectively, of all acceptor sites on these glycoproteins could be galactosylated by this method. |
| |
Keywords: | glycosylation glycosyltransferases galactosyltransferase recombinant glycoproteins enzyme immobilization enzyme reactor |
本文献已被 SpringerLink 等数据库收录! |
|