title: Large-scale production of a soluble human beta-1,4-galactosyltransferase using a Saccharomyces cerevisiae expression system.
abstract: We report in this communication the first large-scale heterologous expression of a glycosyltransferase in yeast. A soluble form of a human beta-1,4-galactosyltransferase (EC 2.4.1.38) was expressed using a Saccharomyces cerevisiae expression system. Fermentation technology afforded the means to increase the expression level of the beta-1,4-galactosyltransferase up to a concentration of 700 mU/liter. The enzyme was produced at a scale of 200 units. The recombinant soluble enzyme was purified 766-fold to a specific activity of approx. 2 U/mg using a purification protocol based on sequential affinity chromatography on N-acetylglucosaminyl- and alpha-lactalbumin-Sepharose, respectively. This study demonstrates that heterologous expression of a glycosyltransferase is possible on a large scale and offers an alternative to natural sources like human breast milk or bovine colostrum.
authors.affiliation: Institute of Biotechnology, Research Center Jülich, Germany.
authors.forename: G F
authors.initials: GF
authors.lastname: Herrmann
authors.forename: C
authors.initials: C
authors.lastname: Krezdorn
authors.forename: M
authors.initials: M
authors.lastname: Malissard
authors.forename: R
authors.initials: R
authors.lastname: Kleene
authors.forename: H
authors.initials: H
authors.lastname: Paschold
authors.forename: D
authors.initials: D
authors.lastname: Weuster-Botz
authors.forename: U
authors.initials: U
authors.lastname: Kragl
authors.forename: E G
authors.initials: EG
authors.lastname: Berger
authors.forename: C
authors.initials: C
authors.lastname: Wandrey
mesh_terms: D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D009237:N-Acetyllactosamine Synthase / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D012995:Solubility
chemical_list: D018076:DNA, Complementary; D011994:Recombinant Proteins; D009237:N-Acetyllactosamine Synthase