Structural and functional analyses of hepatitis B virus X protein BH3-like domain and Bcl-xL interaction.

Hepatitis B virus (HBV) X protein, HBx, interacts with anti-apoptotic Bcl-2 and Bcl-xL proteins through its BH3-like motif to promote HBV replication and cytotoxicity. Here we report the crystal structure of HBx BH3-like motif in complex with Bcl-xL where the BH3-like motif adopts a short α-helix to snuggle into a hydrophobic pocket in Bcl-xL via its noncanonical Trp120 residue and conserved Leu123 residue. This binding pocket is ~2 Å away from the canonical BH3-only binding pocket in structures of Bcl-xL with proapoptotic BH3-only proteins. Mutations altering Trp120 and Leu123 in HBx impair its binding to Bcl-xL in vitro and HBV replication in vivo, confirming the importance of this motif to HBV. A HBx BH3-like peptide, HBx-aa113-135, restores HBV replication from a HBx-null HBV replicon, while a shorter peptide, HBx-aa118-127, inhibits HBV replication. These results provide crucial structural and functional insights into drug designs for inhibiting HBV replication and treating HBV patients.

Prolonged suppression of HBV in mice by a novel antibody that targets a unique epitope on hepatitis B surface antigen.

OBJECTIVE: This study aimed to investigate the therapeutic potential of monoclonal antibody (mAb) against HBV as a novel treatment approach to chronic hepatitis B (CHB) in mouse models. METHODS: Therapeutic effects of mAbs against various epitopes on viral surface protein were evaluated in mice mimicking persistent HBV infection. The immunological mechanisms of mAb-mediated viral clearance were systematically investigated. RESULTS: Among 11 tested mAbs, a novel mAb E6F6 exhibited the most striking therapeutic effects in several HBV-persistent mice. Single-dose administration of E6F6 could profoundly suppress the levels of hepatitis B surface antigen (HBsAg) and HBV DNA for several weeks in HBV-transgenic mice. E6F6 regimen efficiently prevented initial HBV infection, and reduced viral dissemination from infected hepatocytes in human-liver-chimeric mice. E6F6-based immunotherapy facilitated the restoration of anti-HBV T-cell response in hydrodynamic injection (HDI)-based HBV carrier mice. Immunological analyses suggested that the Fcγ receptor-dependent phagocytosis plays a predominant role in E6F6-mediated viral suppression. Molecular analyses suggested that E6F6 recognises an evolutionarily conserved epitope (GPCK(R)TCT) and only forms a smaller antibody-viral particle immune complex with limited interparticle crosslinking when it binds to viral particles. This unique binding characteristic of E6F6 to HBV was possibly associated with its effective in vivo opsonophagocytosis for viral clearance. CONCLUSIONS: These results provided new insight into understanding the therapeutic role and mechanism of antibody against persistent viral infection. The E6F6-like mAbs may provide a novel immunotherapeutic agent against human chronic HBV infection.