Structure-based rational design of peptide inhibitors to disrupt the recognition and interaction between hepatitis B virus large envelope protein and human hepatocyte receptor γ2-adaptin

Abstract

The virion escaping of hepatitis B virus from human hepatocyte is mediated by the recognition and interaction of viral large envelope protein with human hepatocyte receptor γ2-adaptin. In this work, a structure-based rational strategy was described to design peptide inhibitors serving as the interaction disruptors between the preS1 domain of hepatitis B virus large envelope protein and the EAR domain of γ2-adaptin. The binding site of preS1 to γ2-EAR was extracted from the crystal structure of preS1 in complex with EAR as a short preS1^29–36 phage peptide that can extendedly bind on the β-sheet surface of γ2-EAR domain. Computational modeling revealed a systematic amino acid preference profile for the domain–peptide interaction, which was then employed to guide an optimization protocol for improving the biological activity of preS1^29–36 peptide. A number of potential hits were obtained from the evolution, and four were measured in vitro to have a comparable or higher affinity as compared to the native preS1^29–36 peptide. Potent peptide ligands were structurally optimized according to alanine-scanning mutagenesis and complex structural analysis, resulting in the strongest binder with domain-binding affinity at micromolar level. It is revealed that a potent γ2-EAR-binding peptide (8-mer sequence X¹X²X³X⁴X⁵X⁶X⁷X⁸) can be divided into three regions; the region 1 contains residues X² and X³ that should be occupied by negatively charged amino acids such as Glu and Asp, the region 2 covers residues X⁴, X⁵, and X⁶ that directly contact a hydrophobic pocket on γ2-EAR surface, and the region 3 corresponds to residue X⁷ that prefers polar amino acids and is able to form hydrogen bond. In addition, the N-terminal and C-terminal residues X¹ and X⁸ only play a marginal role in the domain–peptide binding.