Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Physicochemical properties of recombinant hepatitis B surface antigen expressed in mammalian cell (C127)

  • 182 Accesses

  • 3 Citations


The physicochemical properties of recombinant hepatitis B surface antigen (r-HBsAg), which was expressed in C127 mammalian cell were studied. Using roller bottle culture in DMEM supplemented with fetal bovine serum, 10–15 mg/L of r-HBsAg was produced with about 31% of purification yield. The purity of r-HBsAG by HPLC was 99.8% and electron microscopic examination showed homogeneous spherical particle with 22 nm in diameter, a morphological characteristic of HBsAg. The density of r-HBsAg by CsCl density gradient method was 1.19 g/ml and the isoelectric point by Mono PTM HR 5/20 column was 4.6. The analysis of subunit protein pattern using SDS-PAGE followed by scanning densitometry gave 81.3% of S protein and 18.7% of pre-S protein. Fluorophore-assisted-carbohydrate-electrophoresis analysis showed the relative amount of carbohydrate to protein was 1.7% and its major component was N-acetyl glucosamine, which was about 39% of total carbohydrate. The relative amount of lipid to protein determined by vanillin phosphoric acid method was 32.5% and its major component was phospholipid, which was about 70% of total lipid. The physicochemical properties of C127 mammalian cell-derived r-HBsAg are similar to those of p-HBsAg, suggesting that the r-HBsAg can be used in developing a new preventive vaccine against hepatitis B.

This is a preview of subscription content, log in to check access.

References Cited

  1. Akahane, Y., Clinical trial of a pre-S2 containing hepatitis B vaccine: Early response of anti pre-S2 in vaccines and efficacy in non-responders to conventional vaccines.International Symposium on Viral Hepatitis and Liver Disease. May 10–14, Tokyo, Japan (1993).

  2. Ansari, G. A. S. and Smith, L., High performance liquid chromatography of cholesterol autooxidation products.J. Chromatogr., 175, 307–315 (1979).

  3. Beasley, R. P. and Hwang, L. Y., Inviral hepatitis and liver disease. Grune & Stratton, New York. pp 209–224 (1984).

  4. Bligh, E. G. and Dyer, W. J., A rapid method of total lipid extraction and purification.Can. J. Biochem. Physiol., 37, 911–917 (1959).

  5. Blumberg, B. S., Alter, H. J. and Visnich, S., A new antigen in leukemia sera.JAMA, 191, 541–546 (1965).

  6. Blumberg, B. S., Sutnick, A. L. and London, W. T., Hepatitis and leukemia: their relation to Australia antigen.Bull. N. Y. Acad. Med., 44, 1566–1586 (1968).

  7. Courouce-Pauty, A. M., Plancon, A. and Soulier, J. P., Distribution of HBsAg subtypes in the world.Vox Sang, 44, 197–211 (1983).

  8. Dane, D. S., Cameron, C. H. and Briggs, M., Virus-like particles in serum of patients with Australia-antigen-associated hepatitis.Lancet, 1, 695–698 (1970).

  9. Diminsky, D., Schirmbeck, R., Reimann, J. and Barenholz, Y., Comparison between hepatitis B surface antigen (HBsAg) particles derived from mammalian cells (CHO) and yeast cells (Hansenula polymorpha): composition, structure and immunogenicity.Vaccine, 15, 637–647 (1997).

  10. Gavilanes, F., Gomez-Gutierrez, J., Aracil, M., Gonzalez-Ros, J. M., Ferragut, J. A., Guerrero, E. and Peterson, D. L., Hepatitis B surface antigen. Role of lipids in maintaining the structural and antigenic properties of protein components.Biochem. J., 265, 857–864 (1990).

  11. Gavilanes, F., Gonzalez-Ros, J. M. and Peterson, D. L., Structure of hepatitis B surface antigen. Characterization of the lipid components and their association with the viral proteins.J. Biol. Chem., 257, 7770–7777 (1982).

  12. Gerlich, W. H., Lu, X. and Heermann, K. H., Studies on the attachment and penetration of hepatitis B virus.J. Hepatol., 17 (Suppl 3), 10–14 (1993).

  13. Heermann, K. H., Goldmann, U., Schwartz, W., Seyffarth, T., Baumgarten, H. and Gerlich, W. H., Large surface proteins of hepatitis B virus containing the pre-S sequence.J. Virol., 52, 396–402 (1984).

  14. Hourvitz, A., Mosseri, R., Solomon, A., Yehezkelli, Y., Atsmon, J., Danon, Y.L., Koren, R. and Shouval, D., Reactogenicity and immunogenicity of a new recombinant hepatitis B vaccine containing Pre S antigens: a preliminary report.J. Viral. Hepats., 3, 37–42 (1996).

  15. Hu, G. F. and Vallee, B. L., A gel retardation assay for the interaction of proteins and carbohydrates by fluorophore-assisted carbohydrate electrophoresis.Anal. Biochem., 218, 185–191 (1994).

  16. Krone, B., Gerlich, W. H., Heermann, K. H., Lenz, A., Lu, X., Seiter, M. and Tomslen, R., Significance of pre-antigens for hepatitis B vaccines.Progress in hepatitis B immunization. 194, 21–25 (1990).

  17. Michel, M. L., Sobczak, E., Malpiece, Y., Tiollais, P. and Streeck, R. E., Expression of amplified hepatitis B virus surface antigen genes in chinese hamster ovary cells.Biotechnology, 3, 561–566 (1985).

  18. Michel, M. L., Pontisso, P., Sobczak, E., Malpiece, Y., Streek, R. E. and Tiollais, P., Synthesis in animal all of hepatitis B surface antigen particles carrying a receptor for polymerized human serum albumin.Proc. Natl. Acad. Sci. USA., 81, 7708–7712 (1984).

  19. Neurath, A. R., Antibodies to a synthetic peptide from the pre-S 120–145 region of the hepatitis B virus envelope are virus neutralizing.Vaccine, 4, 35–37 (1986).

  20. Samanta, H. and Youn, B. W., Expression of hepatitis B virus surface antigen containing the pre-S region in mammalian cell culture system.Vaccine, 7, 69–76 (1989).

  21. Sherlock, S., Fifth international symposium on viral hepatitis summary.J. Hepatol., 17 (Suppl 3), S2-S5 (1993).

  22. Starr, C. M., Masada, R. I., Hague, C., Skop, E. and Klock, J. C., Fluorophore-assisted carbohydrate electro-phoresis in the separation, analysis, and sequencing of carbohydrates.J. Chromatogr. A., 720, 295–321 (1996).

  23. Stibbe, W. and Gerlich, W. H., Structural relationships between minor and major proteins of hepatitis B surface antigen.J. Virol., 46, 626–628 (1983).

  24. Thoma, H. A., Hemmerling, A. E. and Hotzinger, H., Evaluation of immune response in a third generation hepatitis B vaccine containing pre-S proteins in comparative trials.Progress in hepatitis B immunization. 194, 35 (1990).

  25. Tiollais, P., Paucel, C. and Dejean, A., The hepatitis B virus.Nature, 317, 489–495 (1985).

  26. Valenzuela, P., Medina, A., Rutter, W. J., Ammerer, G. and Hall, B. D., Synthesis and assembly of hepatitis B virus surface antigen particles in yeast.Nature, 298, 347–350 (1982).

Download references

Author information

Correspondence to Eung-Chil Choi.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Lee, Y.S., Kim, B.K. & Choi, E. Physicochemical properties of recombinant hepatitis B surface antigen expressed in mammalian cell (C127). Arch. Pharm. Res. 21, 521 (1998). https://doi.org/10.1007/BF02975368

Download citation

Key words

  • Recombinant hepatitis B surface antigen
  • Physicochemical properties
  • Mouse fibroblast cell (C127)
  • Fluorophore-assisted carbohydrate electrophoresis