Glycopeptides as Targets for Dendritic Cells: Exploring MUC1 Glycopeptides Binding Profile toward Macrophage Galactose-Type Lectin (MGL) Orthologs

Abstract

The macrophage galactose-type lectin (MGL) recognizes glycan moieties exposed by pathogens and malignant cells. Particularly, mucin-1 (MUC1) glycoprotein presents an altered glycosylation in several cancers. To estimate the ability of distinct MGL orthologs to recognize aberrant glycan cores in mucins, we applied evanescent-field detection to a versatile MUC1-like glycopeptide microarray platform. Here, as binding was sequence-dependent, we demonstrated that not only sugars but also peptide region impact the recognition of murine MGL1 (mMGL1). In addition, we observed for all three MGL orthologs that divalent glycan presentation increased the binding. To assess the utility of the glycopeptide binders of the MGL orthologs for MGL targeting, we performed uptake assays with fluorescein-MUC1 using murine dendritic cells. A diglycosylated MUC1 peptide was preferentially internalized in an MGL-dependent fashion, thus showing the utility for divalent MGL targeting. These findings may be relevant to a rational design of antitumor vaccines targeting dendritic cells via MGL.

C-Type Lectin Receptor (CLR)–Fc Fusion Proteins As Tools to Screen for Novel CLR/Bacteria Interactions: An Exemplary Study on Preselected Campylobacter jejuni Isolates

“…While two different orthologs (mMGL-1 and mMGL-2) are found in mice, humans only express one MGL isoform (hMGL). It is known that mMGL-2 displays a similar binding profile as hMGL which may account for the marginal binding observed for the mMGL-1 ortholog in our study ( 46 , 47 ).…”

Identification of a secondary binding site in human macrophage galactose-type lectin by microarray studies: Implications for the molecular recognition of its ligands

“…This loss of affinity directly implies that MGL recognizes not only the GalNAc monosaccharide, but also the underlying glycan or protein backbone, through a secondary binding site, of which the His 259 amino acid is a key element. Through MD simulations, we could confirm the MGL-mediated recognition of the peptide backbone in MUC1 (17,22) as well as in MUC2 through the His 259 residue. Strikingly, the type of interactions, either through H bonds or water bridges, depended on the peptide sequence of the glycopeptide.…”

“…Moreover, the underlying Ser of Thr residues in Tn antigen play a key role in the recognition process (20,21). In the case of MGL, additional contacts with the peptide backbone upon binding of MUC1 Tn-glycopeptides have been described previously (17,22). To address whether the His 259 plays a role in the fine specificity of MGL toward Tn-containing glycopeptides, we employed a mucin glycopeptide array that included the known MGL ligands MUC1, MUC2, and CD45, which we interrogated using the MGL short -and MGL short H259T-Fc proteins ( Fig.…”

“…At low concentrations, binding of MGL short was avid for CD45 and MUC2 glycopeptides, whereas the well-known MGL ligand MUC1 (17,22,23) displayed weaker binding to MGL short ( Fig. 2A).…”

Revealing biomedically relevant cell and lectin type-dependent structure–activity profiles for glycoclusters by using tissue sections as an assay platform

“…as free monomeric carbohydrate recognition domain (CRD) and as CRD extended by the a-helical coiled-coil stalk of the transmembrane receptor that confers its trimerisation. Table 1 presents schemes of the quaternary structures and listings of specicity proles (for details on ne-specicity, please see; [43][44][45][46][47][48][49][50][51][52][53][54] for details on structural aspects, please see 48,50,[55][56][57][58][59][60] ). As documented in this table, the relative degrees of affinity to natural GalNAc-containing glycans are not identical.…”