Glycan analysis of monoclonal antibodies secreted in deposition disorders indicates that subsets of plasma cells differentially process IgG glycans.

OBJECTIVE: To compare the glycosylation of polyclonal serum IgG heavy chains in a patient with rheumatoid arthritis (RA) with that of monoclonal serum IgG heavy chains in the same patient during an episode of heavy-chain deposition disease (HCDD), to establish whether glycosylation processing is specific for subsets of B cells. METHODS: Serum IgG was purified using a HiTrap protein G column. Immunoglobulins were run on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels, and IgG glycans were isolated from gel bands and fluorescently labeled. Glycans were analyzed by normal-phase high-performance liquid chromatography and by liquid chromatography-electrospray ionization-mass spectrometry. RESULTS: The glycosylation of serum immunoglobulins from a patient with seronegative RA and HCDD was analyzed. The predominant immunoglobulin was a truncated glycosylated gamma3 heavy chain, and a small amount of polyclonal IgG was also present. The glycan profile showed that the monoclonal gamma3 heavy chain contained fully galactosylated biantennary glycans with significantly less fucose but more sialic acid than in IgG3 from healthy controls. In contrast, the polyclonal IgG showed an RA-like profile, with a predominance of fucosylated biantennary glycans and low levels of galactosylation. The glycan profile of serum IgG obtained from the same patient during disease remission resembled a typical RA profile. CONCLUSION: These data indicate that different types of B cells process a particular set of IgG glycoforms.

Serum amyloid P component in mink, a non-glycosylated protein with affinity for phosphorylethanolamine and phosphorylcholine.

Experimental AA amyloidosis in the mink is used as a model for the amyloid disease process. In that context it is important to characterize the different proteins involved in the amyloid formation. In the present work, we have characterized the serum amyloid P component (SAP) in mink. SAP was purified from serum by affinity chromatography using phosphorylethanolamine-coupled ECH-sepharose 4B. SDS-PAGE showed one major protein band (approximately 26 kDa) together with one minor band (10% of the major band) with a higher molecular mass (approximately 30 kDa) corresponding to a non-glycosylated and a glycosylated variant. All SAP molecules elucidated so far have at least one major subunit that is heavily glycosylated. It is therefore the first time that a non-glycosylated SAP protein is found in a mammalian species. The amino acid sequence was established using Edman degradation and mass spectrometry. As expected, the protein showed high homology with the other mammalian SAP molecules, ranging from 73% (human) to 63% (mouse). The SAP protein showed affinity for phosphorylcholine and thus expressed CRP-like properties.