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. Author manuscript; available in PMC: 2013 Jan 3.
Published in final edited form as: Bioconjug Chem. 2007 Mar 20;18(3):806–814. doi: 10.1021/bc060341n

Figure 6.

Figure 6

Coupling to IgG via N-glycan chains. Selective labeling of the asialo-IgG1 molecule confirmed by PNGase F treatment. (A) Schematic diagram of an IgG molecule with the N-glycan structures attached at the Fc region. (B) The C2 keto galactose moiety was transferred to the carbohydrate in the Fc region of IgG1, using the mutant enzyme β4Gal-T1-Y289L. Mixtures were either treated without PNGase F (−) or with PNGase F (+) containing NP-40, prior to the coupling to aminooxy biotin. The mixtures were resolved by SDS-PAGE, transferred by Western blot to nitrocellulose membranes, and probed with streptavidin–HRP. No chemiluminescence was detected in PNGase F treated sample (+), showing that the transfer of the modified C2 keto galactose moiety is selective for the glycan portion of IgG1 and glycosylation has occurred in the heavy chain of IgG1 at the free GlcNAc residues (10, 25, and 50 ng).