Physicochemical and immunological characterization of hepatitis B virus envelope particles exclusively consisting of the entire L (pre-S1+pre-S2+S) protein

• Published in: Vaccine 2001-04, Vol.19 (23-24), p.3154-3163
• Main Authors: Yamada, Tadanori, Iwabuki, Hidehiko, Kanno, Takashi, Tanaka, Hiroyuki, Kawai, Tomoji, Fukuda, Hideki, Kondo, Akihiko, Seno, Masaharu, Tanizawa, Katsuyuki, Kuroda, Shun'ichi
• Format: Article
• Language: English
• Subjects: AL protein; Animals; Atomic force microscopy; Biological and medical sciences; Chemical Phenomena; Chemistry, Physical; Circular Dichroism; Drug Stability; Fundamental and applied biological sciences. Psychology; Gene Products, env - chemistry; Gene Products, env - immunology; Gene Products, env - isolation & purification; HBsAg L particles; Hepatitis B Antibodies - biosynthesis; Hepatitis B Surface Antigens - chemistry; Hepatitis B Surface Antigens - immunology; Hepatitis B Surface Antigens - isolation & purification; Hepatitis B Vaccines - chemistry; Hepatitis B Vaccines - immunology; Hepatitis B Vaccines - isolation & purification; hepatitis B virus; Hepatitis B virus - chemistry; Hepatitis B virus - immunology; Immune responses; L protein; Mice; Mice, Inbred BALB C; Microbiology; Microscopy, Atomic Force; Pre-S1 region; Receptors, Albumin - chemistry; Receptors, Albumin - immunology; Ultracentrifugation; Vaccines, antisera, therapeutical immunoglobulins and monoclonal antibodies; Virology; Yeast-derived hepatitis B vaccine
• ISSN: 0264-410X; 1873-2518
• DOI: 10.1016/S0264-410X(01)00017-2

The hepatitis B virus (HBV) envelope (env) protein is composed of three regions; the 108- or 119-residue pre-S1 region involved in the direct interaction with hepatocytes, the 55-residue pre-S2 region associated with the polymerized albumin-mediated interaction, and the major 226-residue S protein region. Thus, to improve the immunogenic potency of conventional HB vaccines, development of a new vaccine containing the entire pre-S1 region in addition to pre-S2 and S is desired. We previously reported the efficient production of the HBV env L (pre-S1+pre-S2+S) protein in the recombinant yeast cells [J Biol Chem 267 (1992) 1953]. In this study, the HBV env L protein produced as nano-particles in yeast has been purified and characterized. By equilibrium sedimentation, an average molecular weight of L particle was estimated to be approximately 6.4×106, indicating that about 110 molecules of L proteins are assembled into an L particle. By atomic force microscopy in a moist atmosphere, the L particles were observed as large spherical particles with a diameter of 50–500 nm. The L particles were stable on short-time heating at a high temperature and long-time storage at a low temperature but rather unstable on repeated freezing and thawing and treatment with dithiothreitol. When immunized in mice, L particles elicited efficiently and simultaneously the anti-S, anti-pre-S2, and anti-pre-S1 antibodies. The ED50 values in mice for the anti-S and anti-pre-S2 antibodies were similar to those elicited by the M (pre-S2+S) particles. Furthermore, the anti-pre-S1 rabbit antibodies were found to recognize various segments of the pre-S1 region, including the pre-S1 (21–47) segment. These results show the high ability of L particles to induce all antibodies against HBV env proteins, hence promising the future application of L particles for the next generation HB vaccine.