Synthesis and Microarray-Assisted Binding Studies of Core Xylose and Fucose Containing N-Glycans

Abstract

The synthesis of a collection of 33 xylosylated and core-fucosylated N-glycans found only in nonmammalian organisms such as plants and parasitic helminths has been achieved by employing a highly convergent chemo-enzymatic approach. The influence of these core modifications on the interaction with plant lectins, with the human lectin DC-SIGN (Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Nonintegrin), and with serum antibodies from schistosome-infected individuals was studied. Core xylosylation markedly reduced or completely abolished binding to several mannose-binding plant lectins and to DC-SIGN, a C-type lectin receptor present on antigen presenting cells. Employing the synthetic collection of core-fucosylated and core-xylosylated N-glycans in the context of a larger glycan array including structures lacking these core modifications, we were able to dissect core xylose and core fucose specific antiglycan antibody responses in S. mansoni infection sera, and we observed clear and immunologically relevant differences between children and adult groups infected with this parasite. The work presented here suggests that, quite similar to bisecting N-acetylglucosamine, core xylose distorts the conformation of the unsubstituted glycan, with important implications for the immunogenicity and protein binding properties of complex N-glycans.

How and why plants and human N-glycans are different: Insight from molecular dynamics into the “glycoblocks” architecture of complex carbohydrates

“…Nevertheless, the fact that glycoforms follow recurrent sequence patterns, clearly suggests that the glycans 3D structure is also non-random and very likely sequencedetermined. We use computer modelling to gain insight into these relationships and to define a framework to understand how subtle modifications to the glycans sequence can alter their 3D structure and conformational dynamics, ultimately regulating recognition [19]. In this work we use molecular dynamics (MD) simulations to analyse the effects of the inclusion of motifs typically found in plants and invertebrates N-glycans and immunogenic in mammals [20][21][22][23].…”

On‐Chip Screening of a Glycomimetic Library with C‐Type Lectins Reveals Structural Features Responsible for Preferential Binding of Dectin‐2 over DC‐SIGN/R and Langerin

“…Glycan microarrays were printed as previously described on commercially available NHS‐ester activated glass slides. The immobilization of the ligands was confirmed by incubation with fluorescently labeled lectins of known glycan specificity: plant lectin Concanavalin A (ConA, d ‐Man) and fungal Aleuria aurantia lectin (AAL, l ‐Fuc) .…”

Glycan Arrays: From Basic Biochemical Research to Bioanalytical and Biomedical Applications

“…Glycan modifications can be detected for glycosyltransferases after incubation on slides printed with chemically defined oligosaccharides using fluorescent lectins specific for the transferred sugar (70). With this detection assay, several β(1→2)-core-xylosylated N-glycans were examined as targets for galactosyl-, N-acetylgalactosaminyl-, and fucosyltransferases (71). Only one fucosyltransferase, CeFut6 from Caenorhabditis elegans, was affected by core xylosylation, adding additional information to the profound knowledge about this enzyme's substrate spectrum obtained earlier using glycan arrays and the same detection principle (71,72).…”

“…In one recent exemplary case, this was performed for Ig-fusion proteins of two CLRs, the activating dectin-1 and the ligand-dependent activating/inactivating DC-SIGN (dendritic cell-specific intercellular adhesion molecule 3-grabbing nonintegrin), on microarrays with 153 glucans (14, 83). As human DC-SIGN is involved in human immunodeficiency virus (HIV) infection, the DC-SIGN protein and its murine homologs have been thoroughly studied with glycan arrays (14,18,63,71,93).…”