Chapter 7
Production of Chimeric Hepatitis B Virus Surface Antigens
in Mammalian Cells
Mihaela-Olivia Dobrica, Catalin Lazar, and Norica Branza-Nichita
Abstract
The small (S) envelope protein of the Hepatitis B Virus (HBV), HBV-S, has the unique ability to selfassemble into highly immunogenic subviral particles (SVPs), in the absence of other viral factors, in
eukaryotic cells, including those of nonhepatic origin. This feature is currently exploited for generation
of SVPs exposing heterologous epitopes on their surface that can be used as vaccine candidates to target
various diseases. Here, we describe a simple and robust method for production of such chimeric HBV-S
protein-based SVPs in transiently transfected HEK293T cells and purification from cell supernatants by
ultracentrifugation on sucrose cushion and sucrose step gradients. The SVPs obtained by this methodology
have been successfully used in immunogenicity studies in animal models.
Key words HBV, Antigens, Protein production, Purification, Subviral particles, Vaccine
1 Introduction
A variety of expression systems are available for production of
protein antigens and vaccine development. The ideal vaccine candidate should be highly immunogenic and suitable for high yield
production at low costs. While Escherichia coli is the most costefficient production platform for many soluble proteins, mammalian transmembrane and secretory proteins undergoing complex
folding and posttranslational processing are usually not compatible
with expression in prokaryotic cells [1]. These transformations,
requiring intra- and intermolecular disulfide bonds and specific
glycosylation patterns, are often crucial for the biological properties, including protein antigenicity. By providing the appropriate
eukaryotic milieu for protein processing as well as scalability, yeast
and insect cell cultures have become popular alternative hosts to
prokaryotes for high yield expression of many biopharmaceutical
Blaine A. Pfeifer and Andrew Hill (eds.), Vaccine Delivery Technology: Methods and Protocols, Methods in Molecular Biology,
vol. 2183, https://doi.org/10.1007/978-1-0716-0795-4_7, © The Author(s) 2021
Mihaela-Olivia Dobrica and Catalin Lazar contributed equally to this work.
83
Production of Chimeric Hepatitis B Virus Surface Antigens
in Mammalian Cells
Mihaela-Olivia Dobrica, Catalin Lazar, and Norica Branza-Nichita
Abstract
The small (S) envelope protein of the Hepatitis B Virus (HBV), HBV-S, has the unique ability to selfassemble into highly immunogenic subviral particles (SVPs), in the absence of other viral factors, in
eukaryotic cells, including those of nonhepatic origin. This feature is currently exploited for generation
of SVPs exposing heterologous epitopes on their surface that can be used as vaccine candidates to target
various diseases. Here, we describe a simple and robust method for production of such chimeric HBV-S
protein-based SVPs in transiently transfected HEK293T cells and purification from cell supernatants by
ultracentrifugation on sucrose cushion and sucrose step gradients. The SVPs obtained by this methodology
have been successfully used in immunogenicity studies in animal models.
Key words HBV, Antigens, Protein production, Purification, Subviral particles, Vaccine
1 Introduction
A variety of expression systems are available for production of
protein antigens and vaccine development. The ideal vaccine candidate should be highly immunogenic and suitable for high yield
production at low costs. While Escherichia coli is the most costefficient production platform for many soluble proteins, mammalian transmembrane and secretory proteins undergoing complex
folding and posttranslational processing are usually not compatible
with expression in prokaryotic cells [1]. These transformations,
requiring intra- and intermolecular disulfide bonds and specific
glycosylation patterns, are often crucial for the biological properties, including protein antigenicity. By providing the appropriate
eukaryotic milieu for protein processing as well as scalability, yeast
and insect cell cultures have become popular alternative hosts to
prokaryotes for high yield expression of many biopharmaceutical
Blaine A. Pfeifer and Andrew Hill (eds.), Vaccine Delivery Technology: Methods and Protocols, Methods in Molecular Biology,
vol. 2183, https://doi.org/10.1007/978-1-0716-0795-4_7, © The Author(s) 2021
Mihaela-Olivia Dobrica and Catalin Lazar contributed equally to this work.
83