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Methods Mol Biol. 2018;1776:503-531. doi: 10.1007/978-1-4939-7808-3_33.

Displaying Whole-Chain Proteins on Hepatitis B Virus Capsid-Like Particles.

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Internal Medicine II/Molecular Biology, University Hospital Freiburg, Freiburg, Germany.
Department of Medical Microbiology and Hygiene, Institute of Virology, University Hospital Freiburg, Freiburg, Germany.
Institute of Virology, University Hospital Düsseldorf, Düsseldorf, Germany.
Internal Medicine II/Molecular Biology, University Hospital Freiburg, Freiburg, Germany.


The highly immunogenic icosahedral capsid of hepatitis B virus (HBV) can be exploited as a nanoparticulate display platform for heterologous molecules. Its constituent core protein (HBc) of only ~180 amino acids spontaneously forms capsid-like particles (CLPs) even in E. coli. The immunodominant c/e1 epitope in the center of the HBc primary sequence comprises a solvent-exposed loop that tolerates insertions of flexible peptide sequences yet also of selected whole proteins as long as their 3D structures fit into the two acceptor sites. This constraint is largely overcome in the SplitCore system, where the sequences flanking the loop are expressed as two separate but self-complementing entities, with the foreign sequence fixed to the carrier at one end only. Both the contiguous and the split type of CLP strongly enhance immunogenicity of the displayed sequence but also nonvaccine applications can easily be envisaged. After a brief survey of the basic features of the two HBc carrier forms, we provide conceptual guidelines concerning which foreign proteins are likely to be presentable, or not, on either carrier type. We describe generally applicable protocols for CLP expression in E. coli, cell lysis and CLP enrichment by sucrose gradient velocity sedimentation, plus a simple but meaningful gel electrophoretic assay to assess proper particle formation.


CLP-vaccines; CLPs; Capsid-like particles; HBc; Nanoparticulate antigen carrier; Native agarose gel electrophoresis; Protein-display; SplitCore system; Velocity sedimentation

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