Effects of amino acid substitutions in hepatitis B virus surface protein on virion secretion, antigenicity, HBsAg and viral DNA

Abstract

Background & Aims As important virological markers, serum HBsAg and HBV DNA levels show large fluctuations among chronic hepatitis B (CHB) patients. The aim of this study was to reveal the potential impact and mechanisms of amino acid (AA) substitutions in small hepatitis B surface proteins (SHBs) on serum HBsAg and HBV DNA levels. Methods Serum samples from 230 untreated chronic hepatitis B patients with genotype C HBV were analyzed in terms of HBV DNA levels, serological markers of HBV infection and SHBs sequences. In vitro functional analysis of the identified SHBs mutants was performed. Results Among 230 SHBs sequences, there were 39 (16.96%) sequences with no mutation detected (wild-type, WT) and 191 (83.04%) with single or multiple mutations. SHBs consist of 226 AAs, of which 104 (46.02%) had mutations in our study. Some mutations (e.g. sE2G, sL21R, sG24K, sT47A/K, sC69stop (sC69*), sL95W, sL98V, and sG145R) negatively correlated with serum HBsAg levels. HBsAg and HBV DNA levels from this group of patients had a positive correlation (r = 0.61, p < 0.001). In vitro analysis showed that these mutations reduced extracellular HBsAg and HBV DNA levels by restricting virion secretion and antibody binding capacity. Virion secretion could be rescued for sE2G, sC69*, and sG145R by co-expression of WT HBsAg. Conclusion The serum HBsAg levels were lower in untreated CHB patients with novel SHBs mutations outside the major antigenic region than those without mutations. Underlying mechanisms include impairment of virion secretion and lower binding affinity to antibodies used for HBsAg measurements.

Selection of the highly replicative and partially multidrug resistant rtS78T HBV polymerase mutation during TDF-ETV combination therapy

“…Reduced expression levels of HBsAg because of mutations in the surface protein coding region have been reported previously [20,21]. Accordingly, all three sets of mutations resulted in a significant decrease in surface protein levels (Fig.…”

“…It has been speculated that truncated S proteins, like those having sC69*, may result in secretion incompetent variants due to the deletion of the HBsAg C-terminal domain [21,23]. However, in our study, we could detect HBV-DNA in the supernatant of transfected hepatocytes through PEG precipitation and preS1 or core immunoprecipitation, in secreted particles as well as in exosomes.…”

“…al, have reported on the effects of S mutations on HBV replication and release. They showed that truncated S proteins with sC69* are only detectable intracellularly and through an HA Tag domain fused with HBs [21]. The negative results for the detection of secreted envelop proteins in this study might be due to the internal positioning of the S N-terminal domain (and HA Tag) in secreted particles, which make the epitopes inaccessible to the antibodies.…”

“…In this regard, the emergence of rtS78T/sC69* mutation has been occasionally reported in the literature [21,22,25]. The presence of rtS78T in addition to rtT184S, rtV173L, rtL180M, and rtM204V, was associated with a two-fold reduction in ETV susceptibility; however, in line with our results, the ETV resistance level for this mutation alone did not exceed 1.8-fold of the level of wild type HBV [26–29].…”

Identified OAS3 gene variants associated with coexistence of HBsAg and anti‐HBs in chronic HBV infection

“…Among these, two main hypotheses can be summarized predicated on existing point of views. One hypothesis states that it is associated with mutations in the PreS/S gene, especially the “a” determinant and even the polymerase region . Mutations within the “a” determinant may occur under selective pressure induced by host immunity, antiviral therapy, HBV active or passive immunization .…”