Abstract
Hepatitis B virus (HBV) is a global health burden that causes acute and chronic hepatitis. To develop an HBV-neutralizing antibody that effectively prevents HBV infection, we previously generated a human anti-preS1 monoclonal antibody (1A8) that binds to genotypes A–D and validated its HBV-neutralizing activity in vitro. In the present study, we aimed to determine the fine epitope and paratope of 1A8 to understand the mechanism of HBV neutralization. We performed alanine-scanning mutagenesis on the preS1 (aa 19–34, genotype C) and the heavy (HCDR) and light (LCDR) chain complementarity-determining regions. The 1A8 recognized the three residues (Leu22, Gly23, and Phe25) within the highly conserved receptor-binding motif (NPLGFFP) of the preS1, while four CDR residues of 1A8 were critical in antigen binding. Structural analysis of the epitope–paratope interaction by molecular modeling revealed that Leu100 in the HCDR3, Ala50 in the HCDR2, and Tyr96 in the LCDR3 closely interacted with Leu22, Gly23, and Phe25 of the preS1. Additionally, we found that 1A8 also binds to the receptor-binding motif (NPLGFLP) of infrequently occurring HBV. The results suggest that 1A8 may broadly and effectively block HBV entry and thus have potential as a promising candidate for the prevention and treatment of HBV infection.
Highlights
Published: 7 July 2021Hepatitis B virus (HBV), a small-enveloped DNA virus, infects human hepatocytes and causes acute and chronic hepatitis B, which results in a high risk of developing cirrhosis and hepatocellular carcinoma [1,2]
To determine the fine epitope of 1A8, we substituted each of the residues in the preS1 of genotype C to alanine, constructed a series of expression plasmids of pGST–preS1
The results revealed that the GST–preS1–strep carrying L22A, G23A, or F25A substitution completely lost its binding activity to 1A8 (Figure 2), indicating that the fine epitope of 1A8 is Leu22, Gly23, and Phe25 of the preS1
Summary
Hepatitis B virus (HBV), a small-enveloped DNA virus, infects human hepatocytes and causes acute and chronic hepatitis B, which results in a high risk of developing cirrhosis and hepatocellular carcinoma [1,2]. The number of worldwide chronic carriers in 2016 was estimated to be more than 290 million people [3]. Genotypes A and D are ubiquitous but most prevalent in Africa and Europe, while genotypes B and C are prevalent in Asia and Oceania; genotypes E–H are confined to Asia. Genotypes A–D constitute approximately 90% of total hepatitis B patients [8], whereas genotypes E–H represent less than 5% [8,9]. The HBV envelope contains three membrane proteins—large (L), middle (M), and small (S) proteins [10,11]. The S protein is the common C-terminal domain of the three envelope proteins, the M protein contains an N-terminal preS2 domain and the S domain, and the L protein contains an N-terminal preS1 domain and the preS2 and S domains [11]
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