Introduction
Glycosylation plays multiple important roles in many cellular events ranging from structure to signaling and
recognition, involving protein localization, function and activity in tissues and cells. It is widely known that
glycoprotein analysis requires the identification of glycosylation sites and the glycan structures associated with
each site. In addition to the application of point mutations in traditional biochemistry, the study of glycosylation
sites has also played an important role in the identification of N/O-terminal glycosylation sites. MALDI TOF MS and
nano LC-ESI-MS/MS are commonly used to identify N/O-glycosylation sites in biological samples.
Fig.
1. The standard approaches to determine site-specific glycosylation employing the specific enzymatic proteolysis.
(An H J, et al., 2009)
Our Service
As an advanced protein sequencing and mass spectrometry provider, Creative Proteomics provides you
with a wide range of glycosylation site analysis services to obtain vital information that can be used at every
stage of drug development.
At Creative Proteomics, we offer two glycosylation site analysis services to characterize
glycoproteins.
- A approach to determine site-specific glycosylation is to employ a combination of specific enzymatic proteolysis
(usually with trypsin), fractionation of glycopeptides and glycopeptide analysis by MS.
- Another method is a non-specific protease (usually with pronase) are used for hydrolysis, which is a mixture of
exo- and endo-proteases capable of cleaving essentially any peptide bond, eliminating the disadvantage that
polysaccharide proteins are resistant to trypsin.
Our scientists with extensive experience in glycosylation site analysis have developed a complete glycosylation site
analysis workflow:
Fig. 2. Workflow of our
glycation site analysis services.
Our Technology Platforms
Liquid chromatography-mass spectrometry (LC-MS) was used to determine glycosylation sites as part of peptide mapping
studies. Glycosylation at each individual site was then characterized following release of glycans from each
purified glycopeptide using a range of efficient techniques. Our technology platforms include, but are not limited
to.
- Matrix assisted laser desorption ionization mass spectrometry (MALDI-MS) - Molecular ion data.
- Electrospray mass spectrometry (ES-MS) - Fragment ion information.
- Pulsed amperometric detection (HPAEC-PAD) - Chromatogram of high performance anion exchange chromatography.
- Hydrophilic interaction liquid chromatography (HILIC-FLD) - Fluorescence detection.
Applications
The ICH Q6B guideline requires comprehensive characterization of biopharmaceutical samples glycosylatio. Our
glycosylation site analysis service can help you with the following, but not limited to.
- Identify abnormal glycosylation profiles.
- Determine the occupancy of N/O glycosylation sites.
- Assess the population profile of N-glycans or O-glycans present at each glycosylation site in a glycoprotein.
- Demonstrate batch-to-batch consistency.
- Screen glycoproteins made using different cell lines.
- Meets ICH Q6B requirements for biopharmaceutical characterization and validation for new market applications.
Creative Proteomics is a trusted provider of glycoproteomics analysis services. We offer a wide
range of glycosylation site analysis services to our clients worldwile. Our services guarantee accurate and reliable
results, at quick turnaround time! If you would like more information about specific aspects of our services, feel
free to contact us and we will be happy to answer any questions.
References
- An H J, Froehlich J W, Lebrilla C B. (2009) Determination of glycosylation sites and site-specific heterogeneity
in glycoproteins. Curr Opin Chem Biol. 13(4):421-426.
- Shajahan A, Heiss C, Ishihara M, et al. (2017) Glycomic and glycoproteomic analysis of glycoproteins-a
tutorial. Anal Bioanal Chem. 409(19):4483-4505.
