N-glycopeptide Signatures of IgA2 in Serum from Patients with Hepatitis B Virus-related Liver Diseases

Shu Zhang,Xinyi Cao,Chao Liu,Wei Li,Wenfeng Zeng,Baiwen Li,Hao Chi,Mingqi Liu,Xue Qin,Lingyi Tang,Guoquan Yan,Zefan Ge,Yinkun Liu,Qiang Gao,Haojie Lu

doi:10.1074/mcp.ra119.001722

Abstract

N-glycosylation alteration has been reported in liver diseases. Characterizing N-glycopeptides that correspond to N-glycan structure with specific site information enables better understanding of the molecular pathogenesis of liver damage and cancer. Here, unbiased quantification of N-glycopeptides of a cluster of serum glycoproteins with 40-55 kDa molecular weight (40-kDa band) was investigated in hepatitis B virus (HBV)-related liver diseases. We used an N-glycopeptide method based on 18O/16O C-terminal labeling to obtain 82 comparisons of serum from patients with HBV-related hepatocellular carcinoma (HCC) and liver cirrhosis (LC). Then, multiple reaction monitoring (MRM) was performed to quantify N-glycopeptide relative to the protein content, especially in the healthy donor-HBV-LC-HCC cascade. TPLTAN205ITK (H5N5S1F1) and (H5N4S2F1) corresponding to the glycopeptides of IgA2 were significantly elevated in serum from patients with HBV infection and even higher in HBV-related LC patients, as compared with healthy donor. In contrast, the two glycopeptides of IgA2 fell back down in HBV-related HCC patients. In addition, the variation in the abundance of two glycopeptides was not caused by its protein concentration. The altered N-glycopeptides might be part of a unique glycan signature indicating an IgA-mediated mechanism and providing potential diagnostic clues in HBV-related liver diseases.

Highlights

N-glycosylation is the complex posttranslational modification displayed on many proteins and plays important roles in physiopathological processes
We used an N-glycopeptide method based on 18O/16O C-terminal labeling to obtain 82 comparisons of serum from patients with hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) and liver cirrhosis (LC)
N-glycopeptide of Target 40-kDa Band in LC and HCC—We used an N-glycopeptide method based on 18O/16O C-terminal labeling to obtain comparisons of HBV-related HCC and LC

Summary

Graphical Abstract

18O/16O labeling N-glycopeptide quantification and MRM have been performed to investigate aberrant N-glycopeptides in HBV-related liver diseases. N-glycopeptide Signatures of IgA2 in Serum from Patients with Hepatitis B Virus-related Liver Diseases*□S. Unbiased quantification of N-glycopeptides of a cluster of serum glycoproteins with 40 –55 kDa molecular weight (40-kDa band) was investigated in hepatitis B virus (HBV)-related liver diseases. We used an N-glycopeptide method based on 18O/16O C-terminal labeling to obtain 82 comparisons of serum from patients with HBV-related hepatocellular carcinoma (HCC) and liver cirrhosis (LC). The altered N-glycopeptides might be part of a unique glycan signature indicating an IgAmediated mechanism and providing potential diagnostic clues in HBV-related liver diseases. Serious challenges remain for N-glycopeptide analyses in diseases, such as complexity and diversity of N-glycans [26], and lack of validation It was reported the majority of plasma glycoproteins were 24 glycoproteins, over half of them with the molecular weights of 40 –55 kDa (40-kDa band) [27]. Combination of an 18O/16O labeling N-glycopeptide method and multiple reaction monitoring (MRM) was performed to confirm glycopeptide alterations, which can improve the quantitative power and increase the understanding of their functional impact of the observed changes

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION