Studies on the antigenicity and the structure of the Pre-S2 B cell epitope with synthetic peptides

Abstract

Antigenic properties of the major pre-S2 B-cell epitopes of the hepatitis B virus (HBV) surface antigen (HBsAg) were studied using a pre-S2 specific antiHBsAg monoclonal antibody (H8 mAb), anti-HBsAg polyclonal antibodies and synthetic peptides by competitive ELISA. H8 mAb showed HBV subtype- and peptide size-dependent specificity against synthetic peptides. The HBV subtype-dependent specificity was correlated with a single amino acid substitution at position 141. The peptide, p120-145/adw2, substituted leucine for phenylalanine at the position 141 showed no antigenicity against H8 mAb. The peptide size-dependent specificity was correlated with residues 120/123-129. The truncated peptides excluding these residues, p130-145, showed very low antigenicity compared with p120-145, though the antibody binding site was found to be located in p130-145 by conjugation study. The antigenicity of p130-145 could be increased to the level of p120-145 when p130-145 was conjugated to proteins (bovine serum albumin or ovalbumin) or poly-amino acids (poly-L-glutamic acid or random copolymer of L-glutamic acid and L-alanine). The residues 120/123-129 of p120/123-145 seems to be involved in maintaining the high antigenicity of p120/123-145. In substitution studies of the residues in the sequence p120-129 of p120-145/adr, hydrophilic residues 124-126, STT, and a hydrophobic residue 127, phenylalanine, may play important roles in maintaining high antigenicity. In substitution studies of the residues in the sequence p120-129 of p120-145/adr, p6E-123, the residues 131, 137, 141 and 142, leucine, arginine, phenylalanine and proline, respectively, were identified to involve in the H8 mAb binding, but the residues 132 and 135, leucine and arginine, respectively, did not affect the antigenicity. These results suggest that H8 mAb recognizes a discontinuously conformational epitope included at least residues 131-142. Antigenic properties of the pre-S2 region were also studied using antiHBsA...