Sequencing Glycans

To fully understand the biological relevance of a particular glycan in a glycoconjugate one needs to elucidate the primary sequence and subsequent structure. Although the expression levels of the enzymes needed to synthesize and process glycans (glycosidases and glycosyltransferases) can be measured, the synthetic pathways are not linear and a single compound can be used as a substrate in multiple reactions, making it difficult to accurately determine the reaction’s outcome. Therefore, it becomes impossible to predict the precise sequence and structures of glycans produced by a particular cell.

The first analytical step is typically deglycosylation. Free glycans can be analyzed directly, but they are often derivatized with fluorescent markers such as 2-aminobenzamine (2AB) or with methyl groups (permethylation) to increase the sensitivity of detection, which is essential to observe low abundance glycan species. These modified glycans can be analyzed by mass spectrometry and putative structures can be assigned based on the mass spectral data.

  1. Overview of Glycobiology

    Learn about the core sequences and common modifications of N-linked and O-linked glycans in this video. Analysis of these glycans and/or peptide portions of the glycoprotein can be accomplished with the use of deglycosylation enzymes, which are explained in detail. Unlike other chemical deglycosylation methods, enzymatic treatment is much gentler and can provide complete sugar removal with no protein degradation.

  2. Identification and Characterization of Protein Glycosylation P6039S

    Here we illustrate the use of glycosidases for the analysis of a model glycoprotein: recombinant human chorionic gonadotropin beta (hCGβ), which carries both N-glycans and O-glycans in this video. The technique requires only simple instrumentation and typical consumables, and it can be readily adapted to the analysis of multiple glycoprotein samples.

  3. Elucidating the Complexity of Heparin Oligosaccharide Analysis

    Learn more about the structural elucidation of complex and diverse heparin oligosaccharides using Bacteroides Heparinase I, II and III in combination with downstream mass spec analysis.

FAQs for Sequencing Glycans

Protocols for Sequencing Glycans