FINE MAPPING AND FUNCTIONAL-CHARACTERIZATION OF 2 IMMUNODOMINANT REGIONS FROM THE PRES2 SEQUENCE OF HEPATITIS-B VIRUS

A set of monoclonal antibodies (mAbs) directed against the preS2 regio n of hepatitis B virus (HBV) surface antigen (HBsAg) was generated by immunization of mice with native HBsAg isolated from the blood of HBV carriers. According to (1) mutual competition binding of mAb to natura l HBsAg, (2) recognition of full-length preS2 displayed on hepatitis B core particles, (3) recognition of synthetic partial preS2 peptides, and (4) Western blotting using a fusion protein library of truncated p reS2 fragments of different legths, mAbs were assigned to two groups w hich coincided with groups I and III described by Mimms et al. [Virolo gy 1990; 176: 604-619]. All mAbs recognized linear epitopes and were g lycosylation independent. Six out of eight fins mapped mAbs recognized common epitopes located in the amino-terminal part of the preS sequen ce between amino acids 131 and 144 (group I), and inhibited binding of HBsAg to polymerized human serum albumin. Only two mAbs recognized a carboxy-terminal HBV-genotype-specific epitope covering amino acid res idues 162 to 168 (group III). These mAbs bound to the highly variable proteolysis-sensitive hinge of preS2. Although four out of six mAbs ta rgeted to immunodominant region I require the full-length sequence 131 -L[Q/L]DPRVRGLY[F/L]PAG-144 144, two mAbs recognize the shorter and sl ightly carboxy-terminal-shifted sequences 133-DPRVRGLY[F/L]-141 or 135 -PVRGLY[F/L]PAG-144. Together with previously identified preS2 epitope s 133-DPRVRGL-139, 137-RGLYFPA-143, and 132-QDPR-135, these data indic ate diversity of the immune response against epitopes within the same immunodominant region. This diversity may be generated by a labile sec ondary structure. Sequence analysis suggests the transition from an al pha-helix to a loop structure at this site.