glyco application

Removal of N-Linked & O-Linked Glycans from Glycoproteins

Removal of oligosaccharides from glycoproteins, termed deglycosylation, is often used in order to simplify analysis of the peptide and/or glycan portion of a glycoprotein. Detailed knowledge of the glycan structures helps to correlate them to their respective function. To do this, tools are required for highly sensitive analysis of glycan chains. Both chemical and enzymatic methods exist for removing oligosaccharides from glycoproteins. However, chemical methods such as β-elimination with mild alkali (1) or mild hydrazinolysis (2) often results in the degradation of the protein; whereas, enzymatic methods such as treating with PNGase F (NEB #P0704, NEB #P0705) or Endo H (NEB #P0702, NEB #P0703) (3) for N-glycans are much gentler and can provide complete sugar removal with no protein degradation. Unfortunately, there are no general endoglycosidases that can give complete removal of all O-linked glycans. The broadest specificity O-glycosidase (NEB #P0733, NEB #E0540) is able to cleave only core 1 and core 3 O-glycans (4). Using the O-glycosidase in combination with an appropriate mix of exoglycosidases allows the removal of large O-glycans, after the exoglycosidases have trimmed them to their core. However, for removal of intact O-glycan chains a researcher must still use chemical methods.

Using Glycosidases to Remove, Trim, or Modify Glycans on Therapeutic

 
  1. Kakehi K., et al (1994) J Chromatogr A 680, 209–215. PMID: 7952002
  2. Royle L., et al (2002) Anal Biochem 304, 70–90. PMID: 11969191
  3. Maley, F. et al. (1989) Anal. Biochem., 180, 195-204. PMID: 2510544
  4. Koutsioulis, D. et al (2008) Glycobiology , 18, 799-805. PMID: 18635885

FAQs for Removal of N-Linked & O-Linked Glycans from Glycoproteins

Protocols for Removal of N-Linked & O-Linked Glycans from Glycoproteins

Applications of Deglycosylation

There are a number of reasons to deglycosylate a glycoprotein, including:

  • To remove heterogeneity in glycoproteins for X-ray crystallographic analysis
  • To remove carbohydrate epitopes from antigens
  • To enhance or reduce blood clearance rates of glycoprotein therapeutics
  • To investigate the role of carbohydrates in enzyme activity and solubility
  • To investigate ligand binding
  • For quality control of glycoprotein pharmaceuticals
  • To simplify analysis of the peptide portion of the glycoprotein
  • To simplify the analysis of the glycan component via glycan sequencing

Legal Information

This product is covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB).

While NEB develops and validates its products for various applications, the use of this product may require the buyer to obtain additional third party intellectual property rights for certain applications.

For more information about commercial rights, please contact NEB's Global Business Development team at gbd@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

  1. NEBTV17_720

    NEBTV Episode 17

    Learn about glycobiology and its importance in clinical and diagnostic applications in this episode of NEB TV. Also, hear more about how NEB is setting the bar for product quality in this rapidly growing field.

  2. $name

    Identification and Characterization of Protein Glycosylation P6039S

    Learn how glycosidases are used to analyze multiple glycoprotein samples. Here, the model glycoprotein, hCGβ, which carries both N- and O-glycans, is demonstrated.

  3. Glycobiology_Animation_thumb

    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 can be accomplished with the use of deglycosylation enzymes, which can provide complete sugar removal with no protein degradation.

  4. Heparin_Complexity_thumb

    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.