DEFINITION OF THE PRIMARY STRUCTURE OF HEPATITIS-B VIRUS (HBV) PRE-S HEPATOCYTE BINDING DOMAIN USING RANDOM PEPTIDE LIBRARIES

The pre-S-specific monoclonal antibody MA 18/7 has been shown to inhib it the binding of HBV to HepG2 cells and liver membranes. This antibod y can thus be used to identify the critical residues of the pre-S regi on involved in the hepatocyte-binding domain. Using overlapping 7-mer peptides representing the pre-S region of HBV, the epitope recognized by MA 18/7 was shown to contain sequences from both the pre-S1 and pre -S2 regions, thus indicating that the hepatocyte-binding domain is con formationally dependent. To further characterize tile primary structur e of the hepatocyte-binding domain on the pre-S protein, a phage-displ ayed 15-mer peptide library and a 8-mer solid phase peptide library we re used to analyze the fine specificity of the monoclonal antibody MA 18/7. Several mimotopes were identified with the phage-displayed pepti de library, the majority of which possess a central motif with at leas t three identical residues present within the native pre-S1 sequence. No significant consensus sequences were found when these mimotopes wer e compared to the pre-S2 sequence. Mimotopes identified using the soli d-phase peptide library also contained a similar motif. All phage mimo topes and a single mimotope from the solid-phase peptide library compe ted with recombinant HBsAg particles containing the pre-S1 region for binding to MA 18/7. Mouse antisera raised against four mimotopes from the phage display library reacted with HBsAg particles containing pre- S sequences. The data show that the structure of the pre-S molecule ar ound the conserved DPAF motif in the pre-S region may have a functiona l role in binding HBV to cellular receptors, and that the central moti f identified in mimotopes of this region may offer a novel strategy ta rget for the improvement of existing hepatitis a vaccines which, at pr esent, are mostly devoid of pre-S specificities. (C) 1997 Academic Pre ss.