In a study led by Haojie Lu, PhD, from Fudan University, researchers introduced a new glycoproteomics approach called high-throughput intact glycopeptides quantitation strategy (HTiGQs). They used N,N-dimethylethylenediamine (DMEN) labeling to improve the charge states of the glycopeptides for enhancing their dissociation efficiency in ETD-MS/MS.

Meanwhile, chemical labeling can increase the dissociation of TMT reporter ions in the HCD-MS/MS, achieving a high-throughput quantitation for analysis of site-specific glycosylation. Moreover, the glycoproteomics approach using HTiGQs can quantify 10 samples in a single experiment, greatly reducing the overall LC-MS measurement time. The research team used HTiGQs to analyze the IgG N-glycopeptides from the serum of 90 human patients with varying severities of liver diseases, which is the first systematic study of IgG glycosylation in liver disease at various severities of stages. A total of 313 intact N-glcopeptides from IgG were identified, which represented the largest data set corresponding to the subclass-specific and site-specific N-glycosylation of IgG in human serum reported to date.

They detected dynamic changes in IgG glycosylation during the progression from hepatitis B virus to cirrhosis and hepatocellular carcinoma. And using the glycoproteomics approach they further verified the combination of IgG1-H3N5F1 and IgG4-H4N3 can be used for distinguishing between different stages of liver diseases, which provided important insight for invasive monitoring and precision medicine liver diseases.

Featured Image: The workflow of high-throughput intact glycopeptides quantitation strategy includes protein extract, preparation of TMT and DMEN-labeled glycopeptides, stepped collision energies HCD followed bynproduct-dependent ETD (sceHCD-pd-ETD) acquisition for intact N-glycopeptide identification and quantification and the bioinformation analysis for biomarker discovery, healthy controls (A), HBV patients (B), CIR patients (C), and HCC patients (D). Image: ©Science China Press