Modulation of Glycan Detection on Specific Glycoproteins by Lectin Multimerization

Abstract

Improved methods for studying glycans could spur significant advances in the understanding and application of glycobiology. The use of affinity reagents such as lectins and glycan-binding antibodies is a valuable complement to methods involving mass spectrometry and chromatography. Many lectins, however, are not useful as analytic tools due to low affinity in vitro. As an approach to increasing lectin avidity to targeted glycans, we tested the use of lectin multimerization. Several biotinylated lectins were linked together through streptavidin interactions. The binding of certain lectins for purified glycoproteins and glycoproteins captured directly out of biological solutions was increased using multimerization, resulting in the detection of lower concentrations of glycoprotein than possible using monomeric detection. The analysis of glycoproteins in plasma samples showed that the level of binding enhancement through multimerization was not equivalent across patient samples. Wheat germ agglutinin (WGA) reactive glycans on fibronectin and thrombospondin-5 were preferentially bound by multimers in pancreatic cancer patient samples relative to control samples, suggesting a cancer-associated change in glycan density that could be detected only through lectin multimerization. This strategy could lead to the more sensitive and informative detection of glycans in biological samples and a broader spectrum of lectins that are useful as analytical reagents.

Proteomic identification of mammalian cell surface derived glycosylphosphatidylinositol‐anchored proteins through selective glycan enrichment

“…We speculate that heparan sulfate may sterically interfere with glycan interaction with the lectins used in this study. The performance of lectin affinity capture enrichment in this workflow may be improved by including additional immobilized lectins in the resin mixture whose specificities widen the array of recognized glycan epitopes [57] and/or through the use of lectin multimerization [58]. Unique features of the enrichment methods indicate their suitability for a particular application.…”

Microstructured Block Copolymer Surfaces for Control of Microbe Adhesion and Aggregation

“…These limitations have recently been addressed through synthetic efforts focused on developing materials with clustered oligosaccharides to achieve multivalent, high-avidity binding [ 7 , 8 ]. Other reports have demonstrated that clustering lectins together on surfaces also improves lectin avidity towards complementary glycoproteins, thereby improving detection sensitivities [ 9 ]. Using this methodology, we recently developed cell capture surfaces containing three-dimensional, circular films of a block copolymer, poly(glycidyl methacrylate)- block -4,4-dimethyl-2-vinylazlactone) (PGMA- b -PVDMA).…”

Application of Lectin Microarrays for Biomarker Discovery

“…Single‐step addition may enable enhanced binding through multivalent interactions. Thus, the multimerization method can potentially broaden the range of glycan structures that can be detected and provide more and different information compared to monomeric detection . Because of low background labeling and signal amplification, the indirect labeling methods have advantages of high specificity and sensitivity.…”