Lectin Microarrays for Glycomic Analysis

Abstract

Glycomics is the study of comprehensive structural elucidation and characterization of all glycoforms found in nature and their dynamic spatiotemporal changes that are associated with biological processes. Glycocalyx of mammalian cells actively participate in cell-cell, cell-matrix, and cell-pathogen interactions, which impact embryogenesis, growth and development, homeostasis, infection and immunity, signaling, malignancy, and metabolic disorders. Relative to genomics and proteomics, glycomics is just growing out of infancy with great potential in biomedicine for biomarker discovery, diagnosis, and treatment. However, the immense diversity and complexity of glycan structures and their multiple modes of interactions with proteins pose great challenges for development of analytical tools for delineating structure function relationships and understanding glycocode. Several tools are being developed for glycan profiling based on chromatography, mass spectrometry, glycan microarrays, and glyco-informatics. Lectins, which have long been used in glyco-immunology, printed on a microarray provide a versatile platform for rapid high throughput analysis of glycoforms of biological samples. Herein, we summarize technological advances in lectin microarrays and critically review their impact on glycomics analysis. Challenges remain in terms of expansion to include nonplant derived lectins, standardization for routine clinical use, development of recombinant lectins, and exploration of plant kingdom for discovery of novel lectins.

Pre-B cell receptor binding to galectin-1 modifies galectin-1/carbohydrate affinity to modulate specific galectin-1/glycan lattice interactions

“…Therefore, even though immortalized cells often show alterations of cell surface receptors, pre-BCR relocalization and signalling using Nalm6 and OP9 cells reflect what is observed in primary cells. In lectin microarrays, carbohydrate-binding proteins (lectins; for a list of lectin abbreviations, see Supplementary Table 2) are immobilized onto a solid support at a high spatial density 33,34 . Interrogation of the arrays with fluorescently labelled sample enables elucidation of the glycome based on the distinct carbohydrate-binding specificities of the lectins.…”

A formal approach to the molecular fuzzy lock-and-key

“…Figure 2 shows another example, a frond of the highly flexible 'glycan kelp bed' that coats the cell surface, and, via binding with lectins, triggers even more complicated logical processes. While proteins are constructed from 20 basic amino acids, the glycan kelp bed is formed from as many as 7,000 glycan determinants, and represents a vastly more complex system for information transmission (Cummings, 2009;Gupta et al, 2010). Figure 3, from Dam et al (2007), illustrates a 'bind-andslide' mechanism by which increasing concentration of a lectin species can induce a phase transition topological change.…”

Targeted High-Throughput Glycoproteomics for Glyco-Biomarker Discovery

“…Their abilities to recognize and bind to specific glycans make them ideal for glycan structure specific glycoprotein enrichment. Lectins have been used in biological research as an affinity reagent for the past few decades, with applications such as lectin histochemistry (Brooks et al 1996;Carter and Brooks 2006), lectin blotting (Welinder et al 2009), lectin-affinity chromatography in combination with mass spectrometry (Abbott and Pierce 2010;Zhao et al 2006;Xu et al 2007;Qiu et al 2008;Jung, Cho, and Regnier 2009) and lectin microarray (Gupta, Surolia, and Sampathkumar 2010;Katrlik et al 2010) to examine the glycoproteome of serum and plasma. …”

“…Generally, there are two types of lectin microarrays: the direct assay and reverse-phase dotblot lectin array (Gemeiner et al 2009;Gupta, Surolia, and Sampathkumar 2010). The direct www.intechopen.com assay format immobilizes lectins on a solid surface and applies prelabeled sample over the surface.…”

“…The variations of spotting have been reported to be 10-20% (Kuno et al 2005) and the variation of a reverse-phase dot-blot assay, 10% (Patwa et al 2006), which may be too high to qualify for FDA approval. To preserve the CRD, it has been suggested that glycans of glycosylated lectins may be used as an anchor point for attachment, followed by anchoring to hydroxylamine or hydrazine containing solid surface, which would preserve the CRD of the lectin (Gupta, Surolia, and Sampathkumar 2010). Of course, not all lectins are glycosylated, but this may help lower the variation of a biomarker assay.…”