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Fast NMR-Based Assessment of Cancer-Associated Protein Glycosylations from Serum Samples
Lorena Rudolph1,2, Renia Krellmann1, Darko Castven3
1Institute of Chemistry and Metabolomics, University of Lübeck, Ratzeburger Allee 160, Lübeck 23562, Germany.
Analytical Chemistry
|April 25, 2025
Summary
Nuclear magnetic resonance (NMR) spectroscopy can now detect more N-glycan signals in blood serum and plasma. This advanced method enhances glycoprotein profiling for disease biomarker discovery.
Area of Science:
- Biochemistry
- Analytical Chemistry
- Medical Diagnostics
Background:
- Nuclear magnetic resonance (NMR) spectra of blood serum and plasma contain signals from metabolites, lipoproteins, and N-glycans on acute-phase proteins.
- N-glycan signals, termed glycoprotein A (GlycA) and glycoprotein B (GlycB), are proposed biomarkers for inflammatory and cardiovascular diseases.
- Existing methods like JEDI-NMR are limited to detecting only acetyl signals from N-glycans, missing crucial information from pyranose ring protons.
Purpose of the Study:
- To develop an improved NMR method for comprehensive N-glycan profiling in serum and plasma.
- To enhance the detection of various N-glycan structures, including sialylation, galactosylation, N-acetylglucosaminylation, and fucosylation.
- To enable rapid and robust detection of glycoprotein biomarkers without enzymatic pretreatment.
Main Methods:
- Utilized selective frequency excitation combined with scalar coupling filtering in NMR spectroscopy.
- Applied the method to serum and plasma samples, including isolated glycoproteins and patient samples.
- Compared NMR glycosylation profiles with established UHPLC-MS analysis.
Main Results:
- Significantly increased the number of observable N-glycan signals in NMR spectra.
- Enabled selective detection of sialylation, galactosylation, N-acetylglucosaminylation, fucosylation, and branching complexity of N-glycans.
- Successfully observed sialylated and nonsialylated Lewis antigens, including Lewis a (CA19-9), a known cancer biomarker.
- Demonstrated excellent agreement between NMR glycosylation profiles and UHPLC-MS analysis for isolated glycoproteins.
- Showcased the method's robustness with samples from patients with hepatocellular carcinoma.
Conclusions:
- The developed NMR technique dramatically enhances N-glycan signal detection in serum and plasma.
- This facilitates detailed glycosylation profiling for biomarker discovery, including cancer-associated antigens.
- The method offers a rapid, robust, and enzyme-free approach for glycoprotein analysis in clinical settings.
