What is N glycan profiling?

Glycosylation mainly refers to the enzymatic process that attaches glycans to proteins, lipids, or other organic molecules. MALDI TOF MS and/or UHPLC based techniques determine the overall N-glycan population linked to a purified target molecule or a mixture of molecules. …

What is N glycan analysis?

Protein glycosylation, the addition of single sugars or oligosaccharide chains (glycans) to a peptide backbone, is a common post-translational modification (PTM) that imparts various biological functions (1). …

What does N-glycosylation do?

Protein N-glycosylation is a metabolic process that has been highly conserved in evolution. In all eukaryotes, N-glycosylation is obligatory for viability. It functions by modifying appropriate asparagine residues of proteins with oligosaccharide structures, thus influencing their properties and bioactivities.

What is the N-glycosylation Sequon?

A sequon is a sequence of consecutive amino acids in a protein that can serve as the attachment site to a polysaccharide, frequently an N-linked-Glycan. The sequon for N-glycosylation is either Asn-X-Ser or Asn-X-Thr, where X is any amino acid except proline, Ser denoting serine and Thr threonine.

Is glycan a carbohydrate?

1 Introduction. Glycans, also called polysaccharides, are carbohydrate-based polymers made by all living organisms.

Why is glycan analysis important?

The glycan moieties attached to the proteins can directly affect protein stability, bioactivity, and immunogenicity. Therefore, glycan variants of a glycoprotein product must be adequately analyzed and controlled to ensure product quality.

What is glycan mapping?

The Agilent AdvanceBio Glycan Mapping workflow provides a timesaving, optimized method to generate glycosylation profiles for these important biotherapeutics. The glycan that is initially attached to the protein is then enzymatically modified to generate a wide variety of individual structures (Figure 1).

Is glycosylation good or bad?

Since many biopharmaceutical proteins are glycoproteins in their native state, and proper glycosylation can be critical for their activity, the production of properly glycosylated protein is essential.

Why is N-linked glycosylation so important?

In the ER, N-linked glycosylation serves to ensure proper folding of proteins through the calnexin/calreticulin cycle. Terminally misfolded N-linked glycoproteins are sent for destruction through the ER-associated degradation pathway.

Is asparagine an amino acid?

Asparagine is an amino acid exchange factor.

Why are they called carbohydrates?

Etymology: Carbohydrates are called carbohydrates because the carbon, hydrogen and oxygen they contain are usually in the proportion to form water with the general formula Cn(H2O)n.

Is glycan A sugar?

Glycans are chain-like structures that are composed of single sugar molecules (monosaccharides) linked together by chemical bonds.

How is the N-glycan profile of an antibody determined?

The N-glycan profile of the antibody was analyzed by releasing the N-glycans from the molecule using PNGase F digestion. The released N-glycans were extracted and enriched by solid phase extraction, followed by 2-AB labeling. The labeled N-glycans were then resolved by HILIC chromatography and detected by fluorescence spectroscopy.

Which is the best method for glycan profiling?

A high-analytical sensitivity, high-resolution method for quantitative and qualitative glycan profiling of monoclonal antibodies has been developed. The method enables high confidence identification of even minor glycoforms and isoforms, using UHPLC/FLD/Q-TOF. 2 performance liquid chromatography (UHPLC).

Which is the major species of N glycan?

In Table II, the N-glycan with one terminal galactosylation and fucosylation (G1Fa, b) was the major N-glycan species in both the reference standard and the product formulations at about 50 to 55% of the total N-glycans observed.

How are N-linked glycans contribute to protein folding?

N-linked glycans also contribute to protein folding by steric effects. For example, cysteine residues in the peptide may be temporarily blocked from forming disulfide bonds with other cysteine residues, due to the size of a nearby glycan.