The Glycosylation of Human Serum IgD and IgE and the Accessibility of Identified Oligomannose Structures for Interaction with Mannan-Binding Lectin

Abstract

Analysis of the glycosylation of human serum IgD and IgE indicated that oligomannose structures are present on both Igs. The relative proportion of the oligomannose glycans is consistent with the occupation of one N-linked site on each heavy chain. We evaluated the accessibility of the oligomannose glycans on serum IgD and IgE to mannan-binding lectin (MBL). MBL is a member of the collectin family of proteins, which binds to oligomannose sugars. It has already been established that MBL binds to other members of the Ig family, such as agalactosylated glycoforms of IgG and polymeric IgA. Despite the presence of potential ligands, MBL does not bind to immobilized IgD and IgE. Molecular modeling of glycosylated human IgD Fc suggests that the oligomannose glycans located at Asn354 are inaccessible because the complex glycans at Asn445 block access to the site. On IgE, the additional CH2 hinge domain blocks access to the oligomannose glycans at Asn394 on one H chain by adopting an asymmetrically bent conformation. IgE contains 8.3% Man5GlcNAc2 glycans, which are the trimmed products of the Glc3Man9GlcNAc2 oligomannose precursor. The presence of these structures suggests that the CH2 domain flips between two bent quaternary conformations so that the oligomannose glycans on each chain become accessible for limited trimming to Man5GlcNAc2 during glycan biosynthesis. This is the first study of the glycosylation of human serum IgD and IgE from nonmyeloma proteins.

The DMα and DMβ Chain Cooperate in the Oxidation and Folding of HLA-DM

“…3, B and C). It has been suggested for Igs that the lack of glycan processing in the Golgi is due to steric hindrance (43). However, the DM␣ N165 residue is located on an exposed strand of the Ig domain (44), making it unlikely that it is inaccessible to glycan-modifying enzymes.…”

“…By analyzing DM glycosylation mutants, we can now explain several previous unresolved observations. It has been demonstrated for Igs (43) and the ␣-chains of DR and DM (39,40) that not all N-linked glycans are processed in the Golgi. Here, we have identified glycan DM␣ N165 as the glycan that remains in the highmannose form (Fig.…”

Collectins, C3 complement protein, annexin V and C-reactive protein in acute ischemic stroke: interrelation and implication to upregulated apoptosis and inflammation

“…Concentration of MBL was determined by the earlier described procedure in which ELISA plates were coated with mannan [26] . Concentration of MBL was expressed in g per ml of serum (g/ml).…”

Discrete MBL-MASP Complexes Show Wide Inter-Individual Variability in Concentration: Data from UK vs Armenian Populations

“…MBL purification MBL was isolated from pooled human serum, for use as a quantitative assay standard, as described previously (27)(28). This method involves precipitation of MBL from serum with 7% polyethylene glycol (PEG 3350), binding of MBL-MASP complexes to mannan-agarose, and elution with EDTA-containing buffer.…”

“…Quantitation ofserum MBL by ELISA MBL capable of binding to mannan was measured using an ELISA in which mannan-coated wells were used for capture of MBL (27). Briefly, ELISA plates (Maxisorp, Nunc, Kamstrup, Denmark) were coated with 50~I of I mg/ml mannan (Sigma-Aldrich, M7504) in O.IM Na 2C03 , pH 9.6 (coating buffer).…”