title: In vitro galactosylation of human IgG at 1 kg scale using recombinant galactosyltransferase.
abstract: The Fc effector functions of immunoglobulin G (IgG) antibodies are in part determined by structural features of carbohydrates linked to each of the paired gamma heavy chains in the antibody constant domain (C(H)2). One glycoform that has been shown to be advantageous is G2, where both arms of complex bi-antennary N-glycans terminate in galactose. In vitro treatment with glycosyltransferases can remodel heterogeneous IgG glycoforms, enabling preparation of IgG molecules with homogeneous glycan chains. Here we describe optimization of conditions for use of a soluble recombinant galactosyltransferase in vitro to remodel glycans of human serum IgG, and we demonstrate a scaled-up reaction in which >98% of neutral glycans attached to 1 kg IgG are converted to the G2 glycoform. Removal of glycosylation reagents from the product is achieved in one step by affinity chromatography on immobilized Protein A.
authors.affiliation: Neose Technologies, Inc., 102 Witmer Road, Horsham, Pennsylvania 19044, USA.
authors.forename: Dale
authors.initials: D
authors.lastname: Warnock
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Bai
authors.forename: Katie
authors.initials: K
authors.lastname: Autote
authors.forename: Johnny
authors.initials: J
authors.lastname: Gonzales
authors.forename: Kyle
authors.initials: K
authors.lastname: Kinealy
authors.forename: Boxu
authors.initials: B
authors.lastname: Yan
authors.forename: Jun
authors.initials: J
authors.lastname: Qian
authors.forename: Tom
authors.initials: T
authors.lastname: Stevenson
authors.forename: David
authors.initials: D
authors.lastname: Zopf
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Bayer
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000235:genetics; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002417:Cattle; D005700:Galactosyltransferases / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry* / Q000235:genetics; D007074:Immunoglobulin G / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D025942:Oligosaccharides, Branched-Chain / Q000138:chemical synthesis* / Q000737:chemistry; D011134:Polysaccharides / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics
chemical_list: D000911:Antibodies, Monoclonal; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D025942:Oligosaccharides, Branched-Chain; D011134:Polysaccharides; D011994:Recombinant Proteins; D005700:Galactosyltransferases; C119701:beta-1,4-galactosyltransferase I