Skip to content


You currently have no items in your basket.


Characterization of the Glycome

Glycosylation, the addition of N- or O- linked carbohydrates to proteins, either as a co-translational or post translational modification can be key for the proteins’ interaction with other proteins and it’s own function if folding is affected.Glycosylation is known to be the most common post-translational modification, and more than 50% of all proteins are glycosylated. Not surprisingly, changes in glycosylation have therefore been suggested to be implicated in a number of diseases. (1)

Glycans introduce complexity to the proteins to which they are attached. These modifications vary during the progression of many diseases; thus, they serve as potential biomarkers for disease diagnosis and prognosis. (2) In the field of therapeutic antibodies, the glycosylation of the antibody can fine tune the efficacy of the antibody with changes in glycosylation altering the antibody-dependent cellular cytotoxicity (ADCC) effector function by up to 40 fold, while decreasing Complement-dependent cytotoxicity (CDC) activity. (3)

Branching Glycans

The study of glycosylation has been hindered due to the complexity of these branching structures. There can be a wide range of isotypes whose differing spatial configuration affects the strength of their interaction with receptors.

There are a number of strategies currently used to investigate glycobiology, however these often involve cleaving the glycan from the protein and studying the glycan in isolation either with micro arrays or mass spectrometry which can be expensive and time consuming. Although Mass Spectrometry is a key technique in the field, quickly identifying the weights of the carbohydrates in a system, it does not elucidate the isoforms which may have an impact on the function of the carbohydrate.

Glycosidase and glycosyltransferase enzymes can be specific for different isoforms and these can be used to elucidate some structural characteristics of the glycan. They can be used for a slow construction or destruction of a glycan structure from which inferences can be made.

View Picture Reference

Micro arrays are frequently used to assess which glycans are bound by a protein, or to discover which carbohydrates maybe playing a part in a biological system. They help elucidate the fine specificities of glycan binding proteins. Although knowing this information is useful, there is still much that can be learnt from the starting point of knowing the terminal glycan motifs of the carbohydrate, since these are the parts of the structure free for interaction.

Lectins are a broad class of naturally occurring glycan binding proteins, initially discovered from plant extracts, that had the ability to agglutinate red blood cells. They are a simple and relatively inexpensive way to check for a variety of terminal glycan motifs and can be used in a wide range of assays in much the same way primary antibodies can be, some can also be  used for neuronal tracing, and others have mitogenic properties.

Raising primary antibodies against the carbohydrates themselves is problematic with so many carbohydrate structures being involved in immune signalling pathways when antibody technologies were being initially developed, since glycosylation influences the function of all immune cells. Glycans play a crucial role in intercellular contacts and leukocytes migration. These interactions are important in activation and proliferation of leukocytes and during immune response.(4)

SigLecs are sialic acid binding ImmunoGlobulin-type Lectins – a whole family of proteins expressed by cells of the immune system - that are sensitive to variations in the presentation of sialic acid and it is suggested that different immune responses can be generated based on the Sialic acid linkage detected.  

Lectins can have a number of sugar recognition sites, with the binding affinity increasing with the number of correct matches between the lectin and the carbohydrate, which also relies on the spatial presentation of the sugars. The strength of the binding of a lectin to a sugar can therefore also help to distinguish between different isotypes of carbohydrate that would appear the same on a mass spectrometer. 

Of themselves, lectins are a study area of interest. Some of the well-characterized roles of lectins are in cell-cell communication, cancer metastasis, embryogenesis, host-pathogen interactions, and tissue development (5).

As lectins are purified from natural sources, there can be some variation in the exact affinity of the fine carbohydrate structure; there can be hypervariability in the binding site loops which  generates carbohydrate recognition diversity (6) although the primary carbohydrate preference is secure. Thus, lectin specificity is usually generalised to a broad specificity determined from a sugar monomer that can inhibit lectin binding, or elute the carbohydrate if used with an agarose bound lectin. Further characterisation of lectin binding is determined with micro arrays, sometimes provided by commercial entities and published as at The National Centre for Functional Glycomics. As with antibodies, the use of recombinant technology can help ensure consistency over the long term and recombinant lectins are available in the market.

Other approaches to detecting carbohydrates comes in the form of LectEnz – modified carbohydrate recognising enzymes that retain their binding function but not enzymatic activity.

There are various offerings on the market now that enable a high level overview of glycosylation in an affordable way that can help screen possibilities and direct further studies if more in depth knowledge is required.

CLICK Chemistry

There are a small range of CLICK Chemistry activated carbohydrates that may also help study the glycome further. 

CLICKable monosaccharides are metabolically processed and subsequently incorporated instead of their natural counterparts into glycan structures of proteins .
Step 1: Intracelluar uptake of tetraacetylated CLICKable monosaccharide. Step 2: Deacetylation by endogenous esterases. Step 3: Metabolisation and incorporation into cellular glycan structures. Step 4: Detection of CLICK-functionalized proteins.