A real game changer for glycomics, though, may be the introduction of carbohydrate microarrays to support high-throughput analysis of protein-glycan interactions. These arrays consist of diverse glycans densely attached to a solid surface in an orderly arrangement.
The microarrays would allow simultaneous analysis of multiple glycan-protein interactions using small amounts of carbohydrate samples. While work is still in progress, these microarrays have the potential to supplant ELISA-based microplate array for routine screening of glycan-binding protein.
Since most carbohydrate-binding proteins achieve tight binding through formation of multivalent complexes, both ligand structure and presentation contribute to recognition. Also, since there are many potential combinations of structure, spacing, and orientation to consider, and the optimal one cannot be predicted, high-throughput approaches for analyzing carbohydrate-protein interactions and designing inhibitors are appealing.
In 2010, NIH scientists reported development of a strategy to vary neoglycoprotein density on a surface of a glycan array. This feature was combined with variations in glycan structure and density to produce an array with approximately 600 combinations of glycan structure and presentation. The unique array platform allows the investigators to distinguish between different types of multivalent complexes on the array surface.
The investigators used the technology to rapidly identify potent multivalent inhibitors of Pseudomonas aeruginosa lectin I (PA-IL), a key protein involved in opportunistic infections of P. aeruginosa, and mouse macrophage galactose-type lectin (mMGL-2), a protein expressed on antigen-presenting cells that may be useful as a vaccine-targeting receptor. An advantage of the approach is that structural information about the lectin receptor was not required to obtain a multivalent inhibitor/probe.
The Consortium for Functional Glycomics (CFG), originally funded by the National Institute of General Medical Sciences (NIGMS), provides multiple resources for researchers working in glycomics. Its mission, it says, is to provide a networking forum and glycomics resources that enable investigators to reveal functions of glycans and glycan-binding proteins that impact human health and disease.
The CFG offers glycan microarray screening services, a reagent bank, and free access to its extensive data repositories and molecule databases. In 2006, the CFG teamed up with the Nature Publishing Group (NPG) to create the Functional Glycomics Gateway, which encompassed the existing CFG website and databases and the Functional Glycomics Update from NPG. As of September 1, the Functional Glycomics Gateway became supported solely by the CFG.
One of the resources provided by the CFG is its printed mammalian glycan microarrays that allow glycan-binding proteins to be screened against a wide variety of glycans for binding specificity. To generate the printed array, a growing library of natural and synthetic mammalian glycans with amino linkers is printed onto N-hydroxysuccinimide (NHS)-activated glass microscope slides forming covalent amide linkages. The current mammalian array (version 5.0) has 611 glycan targets.
The discovery that the CFG printed mammalian glycan array can be used to define the specificity of glycan-specific antibodies in human sera opened up the possibility of evaluating the potential of glycan-specific antibodies as biomarkers for cancer. To this end, the CFG assessed the specificity of several commercial and noncommercial glycan-specific mAbs to define their cross-reactivity with other epitopes.
As novel analytical tools that make glycoprotein analysis more accessible are becoming available, the field of functional glycomics is taking shape and growing in prominence among the other omics fields.
Researchers are also refining and extending conventional tools to reveal how glycans behave and work in a functional context. All these technologies will be needed to support both basic research designed to advance the science of glycomics and characterization of the human glycoproteome.