In vitro galactosylation of human IgG at 1 kg scale using recombinant galactosyltransferase.
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.
Warnock
Dale
D
Neose Technologies, Inc., 102 Witmer Road, Horsham, Pennsylvania 19044, USA.
Bai
Xiaomei
X
Autote
Katie
K
Gonzales
Johnny
J
Kinealy
Kyle
K
Yan
Boxu
B
Qian
Jun
J
Stevenson
Tom
T
Zopf
David
D
Bayer
Robert J
RJ
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
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