Probing Blood Plasma Protein Glycosylation with Infrared Spectroscopy

Anal Chem. 2024 Feb 7;96(7):2830-2839. doi: 10.1021/acs.analchem.3c03589. Online ahead of print.

Abstract

The health state of an individual is closely linked to the glycosylation patterns of his or her blood plasma proteins. However, obtaining this information requires cost- and time-efficient analytical methods. We put forward infrared spectroscopy, which allows label-free analysis of protein glycosylation but so far has only been applied to analysis of individual proteins. Although spectral information does not directly provide the molecular structure of the glycans, it is sensitive to changes therein and covers all types of glycosidic linkages. Combining single-step ion exchange chromatography with infrared spectroscopy, we developed a workflow that enables the separation and analysis of major protein classes in blood plasma. Our results demonstrate that infrared spectroscopy can identify different patterns and global levels of glycosylation of intact plasma proteins. To showcase the strengths and limitations of the proposed approach, we compare the glycoforms of human and bovine alpha-1-acid glycoproteins, which exhibit highly variable global levels of glycosylation. To independently evaluate our conclusions, the glycan moieties of human alpha-1-acid glycoprotein were further analyzed using an established glycomics workflow. Importantly, the chromatographic separation of blood plasma improves the detection of aberrant glycoforms of a given protein as compared to infrared spectroscopy of bulk plasma. The presented approach allows a time-efficient comparison of glycosylation patterns of multiple plasma proteins, opening new avenues for biomedical probing.