Modification of Asparagine-Linked Glycan Density for the Design of Hepatitis B Virus Virus-Like Particles with Enhanced Immunogenicity

Abstract

The small envelope proteins (HBsAgS) derived from hepatitis B virus (HBV) represent the antigenic components of the HBV vaccine and are platforms for the delivery of foreign antigenic sequences. To investigate structure-immunogenicity relationships for the design of improved immunization vectors, we have generated biochemically modified virus-like particles (VLPs) exhibiting glycoengineered HBsAgS. For the generation of hypoglycosylated VLPs, the wild-type (WT) HBsAgS N146 glycosylation site was converted to N146Q; for constructing hyperglycosylated VLPs, potential glycosylation sites were introduced in the HBsAgS external loop region at positions T116 and G130 in addition to the WT site. The introduced T116N and G130N sites were utilized as glycosylation anchors resulting in the formation of hyperglycosylated VLPs. Mass spectroscopic analyses showed that the hy-

Predicting HBsAg clearance in genotype A chronic hepatitis B using HBsAg epitope profiling: A biomarker for functional cure

“…The HBsAg is recognised as a highly dynamic conformational protein subunit forming multisubunit particles encompassing an unusually large number of highly conserved conformationally influential cysteine and proline residues throughout the ‘a’ determinant antigenic domain . These factors are thought to influence formation of numerous loop structures and contribute to a structural format which is not readily conducive to resolution …”

“…A 19plex HBsAg epitope mapping assay on a Bioplex ® 200 platform (Figure A), targeting anti‐HBs epitopes across five HBsAg domains has been described previously (Figure B and C). Briefly, epitope display (antigen conformation) and occupancy (‘native’ anti‐HBs recovery) influence the HBsAg profile.…”

“…Briefly, epitope display (antigen conformation) and occupancy (‘native’ anti‐HBs recovery) influence the HBsAg profile. HBsAg profile is reported as fold change (95% CI: ±0.5‐fold) in comparison to the genotype backbone and normalised to the pretreatment profile (Figure and ) …”

“…A 19plex HBsAg epitope mapping assay on a Bioplex ® 200 platform ( Figure 1A), targeting anti-HBs epitopes across five HBsAg domains has been described previously 12 (Figure 1B and C).…”

Maturing Glycoproteomics Technologies Provide Unique Structural Insights into the N-glycoproteome and Its Regulation in Health and Disease

“…These include site-specific analysis of N-glycoproteins isolated to relative purity rather than in complex mixtures (11, 12, 20, 34 -42), N-glycomics analyses of glycans released from glycoproteins (18,38,(43)(44)(45)(46)(47), and identification and quantification of previously glycosylated sites on de-N-glycosylated proteins ("deglycoproteomics") after removal of the entire glycan or with remnant N-glycan core remaining (48 -57). Although these studies per se do not qualify under our definition of glycoproteomics, (site-specific analysis of the glycoproteome at the intact glycopeptide level), they still provide useful information in conjunction with glycoproteomics for the glycobiologist, provided correct experimental design is applied (58).…”