25 HBV infection inhibition by pre-S1 peptides depends on the N-terminal acyl moiety and key amino acids recognised by neutralising antibodies

Abstract

We have previously reported that myristoylated peptides representing an N-terminal preS1-domain of the HBV L-protein block HBV infection of human hepatocytes and HepaRG-cells. Infection interference is due to inactivation of a yet unknown cell-associated factor and requires the preS1 sequence of amino acids 19-48 (subtype ayw). In the present study, we extended our analysis and investigated (1) the role of the N-terminal 18 preS1 amino acids in infection inhibition, (2) the effect of the replacement of the N-terminal myristic acid by other fatty acids (e.g. palmitic acid, cholesterol and various saturated and unsaturated acyl residues) and (3) the importance of key amino acids within preS1-epitopes that are recognised by the neutralising antibodies MA18/7 (aa 20-23) and 5a19 (aa.26-32). Our results show that although inactive on their own, amino acids 2-19 are essential for infection interference by preS2-48myr, indicating a possible spacer function. Exchange of the N-terminal myristic acid by other hydrophobic residues did not abrogate infection inhibition, indicating no absolute requirement for this fatty acid. Moreover, the introduction of short deletions and single amino acids exchanges within preS19-48 allowed us to functionally map key amino acids required for inhibition. Our data shed light on the molecular mechanism of HBV entry into hepatocytes, as well as providing the possibility of modulating a peptide’s specific activities (including an increase of the ID50 to picomolar levels). Furthermore, these studies provide the basis for the development of new hepadnaviral entry inhibitors for therapeutic approaches.