O-glycosylated HBsAg peptide can induce specific antibody neutralizing HBV infection

Biochim Biophys Acta Gen Subj. 2022 Jan;1866(1):130020. doi: 10.1016/j.bbagen.2021.130020. Epub 2021 Sep 25.


Background: Hepatitis B virus (HBV), which causes hepatitis, liver cirrhosis, and hepatocellular carcinoma, is a global human health problem. HBV contains three envelope proteins, S-, M-, and L-hepatitis B surface antigen (HBsAg). We recently found that O-glycosylated M-HBsAg, reactive with jacalin lectin, is one of the primary components of HBV DNA-containing virus particles. Thus, we aimed to analyze and target the glycosylation of HBsAg.

Methods: HBsAg prepared from the serum of Japanese patients with HBV were analyzed using mass spectrometry. The glycopeptide modified with O-glycan was generated and used for immunization. The specificity of the generated antibody and the HBV infection inhibition activity was examined.

Results: Mass spectrometry analysis revealed that T37 and/or T38 on M-HBsAg of genotype C were modulated by ±NeuAc(α2,3)Gal(β1,3)GalNAc. Chemically and enzymatically synthesized O-glycosylated peptide (Glyco-PS2) induced antibodies that recognize mainly PreS2 in M-HBsAg not in L-HBsAg, whereas the non-glycosylated peptide (PS2) induced antisera recognizing L-HBsAg but not O-glycosylated M-HBsAg. The removal of O-glycan from M-HBsAg partly decreased the reactivity of the Glyco-PS2 antibody, suggesting that peptide part was also recognized by the antibody. The antibody further demonstrated the inhibition of HBV infection in human hepatic cells in vitro.

Conclusions: Glycosylation of HBsAg occurs differently in different HBsAgs in a site-specific manner. The new Glyco-PS2 antibody, recognizing O-glycosylated M-HBsAg of genotype C, could inhibit HBV infection.

General significance: The detailed analysis of HBsAg identified different glycosylations of HBV surface. The glycosylated peptide based on mass spectrometry analysis showed higher potential to induce functional antibody against HBV.

Keywords: Antibody; Glycopeptide; HBsAg; Hepatitis B virus; O-glycan.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies / immunology
  • Antibodies, Neutralizing / immunology
  • Cell Line, Tumor
  • Glycosylation
  • Hep G2 Cells
  • Hepatitis B / immunology*
  • Hepatitis B Surface Antigens / immunology*
  • Hepatitis B Surface Antigens / metabolism*
  • Hepatitis B virus / immunology
  • Hepatitis B virus / pathogenicity
  • Humans
  • Liver / metabolism
  • Peptides / immunology


  • Antibodies
  • Antibodies, Neutralizing
  • Hepatitis B Surface Antigens
  • Peptides