Glycans chemical methods

The lack of a pan-specific or broad specificity O-glycanase makes chemical methods the most appropriate approach for the non-specific release of intact O-linked oligosaccharides from glycoproteins for site analysis.219 The most common chemical methods employed for the release of O-linked glycans are based on (3-elimination. The methods were originally developed to release the glycans for... 

Biosynthesis of GPI anchors starts with the core structure assembly by sequential addition of UDP-GlcNAc (followed by iV-deacetylation), dolichol-phosphate-mannose, and phospho-ethanolamine to phosphatidylinositol and culminates in the en bloc transfer to protein shortly after the protein is synthesized. However, the biosynthetic pathways can differ strikingly between different organisms with respect to specific modifications and fatty acid remodeling occurring after completion of the core glycan. This also applies for the point when certain modifications are introduced, e. g. before or after the transfer of the GPI-moiety to the protein. GPI anchors can be cleaved at defined positions by an array of enzymatic and chemical methods, respectively (O Fig. 5). Thus, it becomes possible to identify GPI-anchored proteins and, moreover, analyze the structure and biosynthesis of GPI anchors [103]. 

Fig. 2. Structures of glycans found in Dictyostelium discoideum. Structures of known carbohydrate components in Dictyostelium discoideum. The structures have been determined by physical/chemical methods or by the analysis of radiolabeled materials. (A) Lipid-linked oligosaccharide precursor of N-linked oligosaccharides [14] (B) truncated lipid-linked oligosaccharide precursor found in mutant strains HL24I and HL243[I4] (C) oligosaccharide with GlcNAc residues at the intersecting position and locations of the methylphosphate esters [16].

N-linked glycans may be released with chemical cleavage (e.g., alkaline-based S-elimination, hydrolysis with hydrazine, or trifluoromethansulfonic acid (TFMS). Some method optimization will be needed, and care must be taken to avoid chemical destruction of the reducing end. Hydrazinolysis, introduced in the 1960s, is the most commonly used chemical method of choice though the harshness of the method often leads to chemical deacetylation of sialic acids and N-acetylhexosamines. Nearly all methods will result in a reactive aldehyde reducing end that can be derivatizated (see below) with the appropriate amine containing molecules. 

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