Raju “Glycosylation Variations with Expression Systems and their impact on biological activity of therapeutic immunoglobulins” BioProcess International, April 2003

Fig. 4 above shows the largest N-linked oligosaccharide structure found in human IgG. This structure is present as a very minor component. The majority of oligosaccharides found in human IgG are truncated versions of the structure shown in Fig. 4. The microheterogeneity of human IgG glycans is mainly due to the presence and/or the absence of eitehr core fucose, bisecting GlcNAc, termial sialic acid, exposed Gal, and often the exposed GlcNAc residues. See Raju above 

In human IgG1, which is the main subtype used for therapeutics, the majority of the Fc glycans are complex biantennary structures wtih variable galactosylation: 1, 1, and 2 terminal galactoses corresponding to G0, G1 and G2 glycoforms, respectively. (Boeggeman, Bioconjugate Chem. 2009, 20, 1228-1236).

G0 glycan: refers to the complex N-linked glycan having the G1cNAc2Man3G1cNAc2 core structure, where no terminal sialic acids (NeuAcs) or terminal galactose (Gal) sugar residues are present. In IgG-GO glycoforms, neither arm is galactosylated so that a GlcNAc residue is exposed at the terminals of each arm. About 20% of the IgG glycans at this glycosylation site terminate in GlcNAc on both arms The IgG-G0 glycoforms have been shown to bind MBL. (Arnold, J. Immunol, 2004 173: 6831-6840). 

G1 glycan: refers to the complex GNGN biantennary N-glycan having the G1cNAc2Man3G1cNAc2 core structure plus one terminal galactose residue. In IgG-G1 glycoforms, a terminal galactose residue is missing from one arm, exposing a GlcNAc residue. (Arnold, J. Immunol, 2004 173: 6831-6840)

G2 glycan: refers to complex GNGN biantennary N-glycan having the G1cNAc2Man3G1cNAc2 core structure plus two terminal galactose residues. (Arnold, J. Immunol, 2004 173: 6831-6840) 

Production of Immunoglobuilin Glycoforms

G0 glycoforms: Raju (US2007/0041979) discloses preparation of G0 glycoform by treating IgG samples with a sialidase A to remove minor amounts of terminal sialic acid residues followed by treating with beta galactosidase to remove terminal beta-galactose residues. After purificaiton on a protein A column, the resulting G0 glycoform was characterized by MALDI-TOF-MS.

Fox (US2010/0266607) discloses an antibody glycoform in which sialic acid and terminal beta galactose residues are removed by incubation with sialidase and beta galactosidase.

G1 glycoforms: (Hueller, US14348822) discloses a method for producing an immunoglobuilin with a G1 glycostructure from a mixture of a glycosylated antibody or fusion polypeptides of the G0, G1 and G2 glycostructures. The method involves the following sequences of events 1) incubate with a galactosyltransferase, 2) incubate this product with a sialytransferase, 3) incubate this product with a beta-1,4-galactosidase to oobtain a beta-1,4-galactosidase reaction product, 4) remove or inactive the beta-1,4-galactosidase and 5) incubate the reaction product with a sialidase. 

G2 glycoforms: Raju (US2007/0041979) discloses preparation of G2 glycoform by treating IgG samples with a sialidase A, purifying on Protein A and then treating the sialidase A IgG samples with beta 1, 4 GT (including UDP-Gal) for 24 hours.

Fox (US2010/0266607) discloses an antibody glycoform which is maximally sialyated by incubation of antibody with beta 1,4-galactosyltransferase and alpha 2,3-sialytransferase and UDP-galactose, CMP-N-acetylnueraminic acid and MnCl.

Free of Core Fucoses: (Wong, US2011/0263828) discloses methods for modifying the Fc region glycosylation pattern of an antiboyd by providing a clycosylated Fc region, contacting the glycosylated Fc region with alpha-fucosidase to remove core fucose residues, contacting the de-fuosylated Fc with beta-1,4-galactosyltransferaseand then alpha-2,6-sialytransferase, then with beta-1,4 galactosyltransferase under conditions suitable for attaching a galatose to the oligosaccharide and then contacting the Fc region with alpha-2,6-sialytransferaseunder conditions suitable for attaching a terminal sialyl residue. 

Free of Galactose:

Raju (US2010/0260751) discloses that host cells may be devoicd of galactosyltransferases such that antiboides expressed in these cells lack galactose. Wihtout galactose, sialic acid will also not be attached.

–Removal of (alpha 1,3) glycosidically bound terminal galactose residues):  diclsoes that alpha 1,3 galactosidases from green coffee beans (EC 3.2.1.22) can be used to selectively remove (alpha 1,3) glycosidically bound terminal galactose residues from the oligosaccharide attached at amino acid residue Asn297 to an immunoglobuiolin CH2 domain.