Raju, T. S., Briggs, J. B., Borge, S. M. & Jones, A. J. Species-specific variation in glycosylation of IgG: evidence for the species-specific sialylation and branch-specific galactosylation and importance for engineering recombinant glycoprotein therapeutics. Glycobiology 10, 477-486

Analytical Chemistry, Genentech Inc., 1 DNA Way, South San Francisco, CA 94080, USA.
Glycobiology (Impact Factor: 3.15). 06/2000; 10(5):477-86. DOI: 10.1093/glycob/10.5.477
Source: PubMed


Immunoglobulins (IgG) are soluble serum glycoproteins in which the oligosaccharides play significant roles in the bioactivity and pharmacokinetics. Recombinant immuno-globulins (rIgG) produced in different host cells by recombinant DNA technology are becoming major therapeutic agents to treat life threatening diseases such as cancer. Since glycosylation is cell type specific, rIgGs produced in different host cells contain different patterns of oligosaccharides which could affect the biological functions. In order to determine the extent of this variation N-linked oligosaccharide structures present in the IgGs of different animal species were characterized. IgGs of human, rhesus, dog, cow, guinea pig, sheep, goat, horse, rat, mouse, rabbit, cat, and chicken were treated with peptide-N-glycosidase-F (PNGase F) and the oligosaccharides analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) for neutral and acidic oligosaccharides, in positive and negative ion modes, respectively. The data show that for neutral oligosaccharides, the proportions of terminal Gal, core Fuc and/or bisecting GlcNAc containing oligosaccharides vary from species to species; for sialylated oligosaccharides in the negative mode MALDI-TOF-MS show that human and chicken IgG contain oligosaccharides with N-acetylneuraminic acid (NANA), whereas rhesus, cow, sheep, goat, horse, and mouse IgGs contain oligosaccharides with N-glycolylneuraminic acid (NGNA). In contrast, IgGs from dog, guinea pig, rat, and rabbit contain both NANA and NGNA. Further, the PNGase F released oligosaccharides were derivatized with 9-aminopyrene 1,4,6-trisulfonic acid (APTS) and analyzed by capillary electrophoresis with laser induced fluorescence detection (CE-LIF). The CE-LIF results indicate that the proportion of the two isomers of monogalactosylated, biantennary, complex oligosaccharides vary significantly, suggesting that the branch specificity of beta1, 4-galactosyltransferase might be different in different species. These results show that the glycosylation of IgGs is species-specific, and reveal the necessity for appropriate cell line selection to express rIgGs for human therapy. The results of this study are useful for people working in the transgenic area.

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Available from: T. Shantha Raju, Dec 30, 2013
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    • "Different explanations have been proposed for this effect: (i) ammonia induces the synthesis of UDP-GlcNAc that competes with the transport of cytosine monophosphate (CMP)–sialic acid to the cell membrane thus inhibiting the sialylation process (Valley et al., 1999) and (ii) ammonia is correlated with the sialylation level through the resulting changes in internal pH in the Golgi apparatus which causes lower activity of glycosyltransferase enzymes (Andersen and Goochee, 1994; Yang and Butler, 2002). However, the reports regarding the effects of ammonia and pH on glycosylation are sometimes contradictory and may be highly dependent on the cell line, cultivation process and glycoprotein type (Raju et al., 2000; Butler, 2006; Chen and Harcum, 2006). For example, changes in sialylation of recombinant human granulocyte colony-stimulating factor (Andersen and Goochee, 1994) have been reported for either low ammonia levels of 2 mM or for very high ammonia levels above 9 mM for other glycoproteins (Chen and Harcum, 2006; Yang and Butler, 2000; Borys et al., 1993). "
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    • "N-glycosylation in animals is a largely cell-type and species-specific feature (Raju et al., 2000; Croset et al., 2012). Moreover, potential glycosylation sites on a given protein can be either unmodified or occupied by varying glycan structures which result from the maturation of the glycan throughout the endoplasmic reticulum (ER) and the Golgi apparatus (GA), leading to microheterogeneity of glycoproteins (Kolarich et al., 2012). "
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