Simultaneous expression of the S and L surface antigens of hepatitis B, and formation of mixed particles in the methylotrophic yeast, Hansenula polymorpha

• Published in: Yeast (Chichester, England) Vol. 7; no. 5; p. 431
• Main Authors: Janowicz, Z A, Melber, K, Merckelbach, A, Jacobs, E, Harford, N, Comberbach, M, Hollenberg, C P
• Format: Journal Article
• Language: English
• Published: England 01.07.1991
• Subjects: Base Sequence; Blotting, Western; Culture Media; DNA, Fungal - chemistry; Fermentation; Gene Expression Regulation, Fungal; Genetic Vectors; Hepatitis B Surface Antigens - biosynthesis; Hepatitis B Surface Antigens - genetics; Hepatitis B Surface Antigens - immunology; Hepatitis B virus - genetics; Hepatitis B virus - immunology; Immunoblotting; Molecular Sequence Data; Pichia - genetics; Pichia - metabolism; Plasmids; Promoter Regions, Genetic; Protein Precursors - biosynthesis; Protein Precursors - genetics; Protein Precursors - immunology; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Recombinant Proteins - immunology; Restriction Mapping
• ISSN: 0749-503X
• DOI: 10.1002/yea.320070502

An expression system has been developed for the methylotrophic yeast Hansenula polymorpha and used to co-express both the L (preS1-S2-S) and S hepatitis B surface antigens (HBsAg) under the control of strong methanol-inducible promoters derived from the methanol oxidase and from the formate dehydrogenase genes. A unique feature of this H. polymorpha expression system is the possibility of integrating up to 100 copies of an expression cassette via a multimeric integration mechanism. Several multimeric integrants containing various numbers of L and S expression cassettes were constructed to give a spectrum of strains characterized by different L to S ratios. The expression level of S antigen was 5-8% of the total soluble cell protein. Analysis by sucrose and CsCl density gradient centrifugation and by particle-specific immunoassays demonstrated that the synthesized HBsAg spontaneously assembled into composite subviral particles containing both S and L proteins. Only a minor portion of the L protein was found to be glycosylated. These H polymorpha-derived composite particles can be used for the production of a hepatitis B virus vaccine with the potential for improved immunogenicity due to the presence of a wider spectrum of epitopes and negligible glycosylation.