Dataset Information

High throughput quantitative analysis of site-specific N-glycosylation reveals glyco-signatures for diagnosis of liver disease

ABSTRACT: Glycosylation influences diverse life processes and changes in pathological processes. The glycoproteomic has been recognized as a hot area to screen markers as altered glycosylation is a universal feature of cancer cells. However, the high throughput screening of glycosylation markers from a large number of clinical samples faces greater technical challenges than peptides without or with other modifications. The most prominent problem in tandem mass tag-based high throughput quantification of intact glycopeptides is the inefficient cleavage of peptide backbone, glycosidic bonds and tandem mass tag concurrently to offer successful peptide and glycan composition determination as well as good quantification. Here, we managed to implement the tandem mass tag labeling into quantitative glycoproteomics by developing a chemical labeling-assisted complementary dissociation method for multiplexed analysis of intact N-glycopeptides. Chemical labeling of intact N-glycopeptides with charge-increasing reagent improved the efficiency of electron transfer dissociation (ETD) significantly for their identification and also increased the efficiency of higher-energy collision dissociation (HCD) in generating reporter ion for quantification, thereby capitalizing on the complementary nature of these two dissociation methods in quantification of intact N-glycopeptides. By using this new strategy, we investigated the site-specific glycosylation of IgG collected from 90 human serum including three stages of liver diseases, hepatitis B virus (HBV), cirrhosis (CIR), hepatocellular carcinoma (HCC) and healthy controls. As a result, 313 intact N-glycopeptides from IgG were identified in total, which represents the most comprehensive site-specific and subclass-specific N-glycosylation of human serum IgG to date. Quantitative glycoproteomics revealed that the combination of IgG1-H3N5F1 and IgG4-H3N4 displayed powerful prediction capability for distinguishing different stages of liver disease patients, which has potential for non-invasive monitoring and pre-stratiﬁcation of liver disease diagnosis. Moreover, we analyzed the human serum using this newly developed method and identified near 2000 intact N-glycopeptides from only 10 L serum and showed this method has applicability for quantification of intact N-glycopeptides in different studies.

ORGANISM(S): Homo Sapiens

SUBMITTER: Haojie Lu

PROVIDER: PXD027379 | iProX | Fri Jul 16 00:00:00 BST 2021

REPOSITORIES: iProX

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High-throughput site-specific <i>N</i>-glycoproteomics reveals glyco-signatures for liver disease diagnosis.

Sun Zhenyu Z Fu Bin B Wang Guoli G Zhang Lei L Xu Ruofan R Zhang Ying Y Lu Haojie H

National science review 20220405 1

The glycoproteome has emerged as a prominent target for screening biomarkers, as altered glycosylation is a hallmark of cancer cells. In this work, we incorporated tandem mass tag labeling into quantitative glycoproteomics by developing a chemical labeling-assisted complementary dissociation method for the multiplexed analysis of intact <i>N</i>-glycopeptides. Benefiting from the complementary nature of two different mass spectrometry dissociation methods for identification and multiplex labelin ...[more]

PMID: 36879659

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Project description:Changes in IgG glycosylation have been reported to be associated with colorectal cancer (CRC), but the alteration in specific subclass of IgG is unknown. Herein, we optimized five common IgG glycopeptides enrichment methods to acquire comprehensive profiling of IgG glycoproteomics in CRC. Notably, incomplete tryptic digestion of IgG occurred when using ordinary ratio of protease to protein, which significantly impacted the final statistical analysis of association between IgG glycosylation and CRC. We introduced a method based on two-step enzymatic digestion, enabling the complete digestion of IgG glycopeptides and further improving the detection intensity of the target glycopeptides. By this approach, total 47 IgG glycopeptides can be identified by nanoLC-ESI-MS/MS. Following rapid and automatic data processing through the self-developed program, we observed that IgG1_H3N4F1 and IgG1_H3N4 were substantially increased in CRC, while IgG1_H5N5F1, IgG1_H5N4F1S1and IgG2_H5N4F1 were markedly decreased. Strikingly, these four glycopeptides were continually significantly changed in early and late stages of CRC. Further evaluation of the diagnostic performance showed they all obtained a fair performance in discriminating the patients from the normal. In terms of the glycan features, it demonstrated that CRC progression associates with the increased agalactosylation, and the decreased digalactosylation and galactosylation per antenna on diantennay glycans of IgG1 and IgG2. Concurrently, the decreased sialylation of IgG1 was strongly correlated with CRC. Moreover, tumor-specific glycosylation analysis showed that the alterations of subclass-specific IgG glycosylation were more significant in colon cancer, and no obvious difference between colon and rectal cancer was found. More importantly, all these findings were validated in an independent cohort. This two-step enzymatic approach is crucial to explore the relationship between the disease progression and IgG glycosylation.

Dataset Information

High throughput quantitative analysis of site-specific N-glycosylation reveals glyco-signatures for diagnosis of liver disease

Publications

High-throughput site-specific <i>N</i>-glycoproteomics reveals glyco-signatures for liver disease diagnosis.

Similar Datasets

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