In Saccharomyces cerevisiae, we synthesized and secreted L-HBVsAg (named as pre-S(Met1 to Asn174)::S(Met175 to lle400)) and three mutants, i.e., pre-S degrees degrees::S (Asn15Gln and Asn123Gln), pre-S degrees degrees::S degrees (Asn15Gln, Asn123Gln, and Asn320Gln), and pre-S degrees degrees::S degrees degrees (Asn15Gln, Asn123Gln, Asn233Gln, and Asn320Gln). All of the secreted pre-S::S was N-glycosylated, i.e., hypermannosylated. In the secretion of pre-S degrees degrees::S degrees and preS degrees::S degrees, besides the hyper-mannosylated form, another immunoreactive protein with much lower molecular mass was observed, which seems to be unglycosylated form of pre-S degrees degrees::S and pre-S degrees degrees::S degrees. Only a part of the secreted pre-S"::S or pre-S"::S' molecules was N-glycosylated, and the site for the partial N-glycosylation seems to be Asn233 in S-antigen region. Compared to the N-glycosylated pre-S degrees degrees::S and pre-S degrees degrees::S degrees, pre-S degrees degrees::S degrees degrees (non-N-glycosylated mutant) was secreted with lower secretion efficiency but showed apparent immunoreactivity to anti-S antigen monoclonal Ab. Interestingly, unlike pre-S degrees degrees::S degrees degrees with authentic C-terminus, the recombinant pre-S degrees degrees::S degrees degrees with C-terminal myc or poly-histidine tag (pre-S degrees degrees::S degrees degrees::tag) was almost all aggregated into insoluble proteins in the intracellular region. Conclusively, the C-terminal sequence and glycosylation in S-antigen region seem to be of crucial importance in determining the secretion efficiency of L-HBVsAg in S. cerevisiae. (c) 2005 Wiley Periodicals, Inc.