NMR assignments of the N-glycans of the Fc fragment of mouse immunoglobulin G2b glycoprotein

The Fc portion of immunoglobulin G (IgG) promotes defensive effector functions in the immune system by interacting with Fc gamma receptors and complement component C1q. These interactions critically depend on N-glycosylation at Asn297 of each C(H)2 domain, where biantennary complex-type oligosaccharides contain microheterogeneities resulting primarily from the presence or absence of non-reducing terminal galactose residues. Crystal structures of Fc have shown that a pair of N-glycans is located between the two C(H)2 domains. Here we applied our metabolic isotope labeling technique using mammalian cells for in-solution structural characterization of mouse IgG2b-Fc glycoforms with a molecular mass of 54 kDa. Based on spectral assignments of the N-glycans as well as polypeptide backbones of Fc, we probed conformational perturbations of Fc induced by N-glycan trimming, especially enzymatic degalactosylation. The results indicated that degalactosylation structurally perturbed the Fc region through rearrangement of glycan-protein interactions. The spectral assignments of IgG2b-Fc glycoprotein will provide the basis for NMR investigation of its dynamic conformations and interactions with effector molecules in solution.

Automated summary

The N-glycosylation at the Fc portion of IgG plays a critical role in defensive effector functions. Degalactosylation was found to structurally perturb the Fc region in a solution-based analysis.