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Title: Expression of hepatitis B virus chimeric proteins in prokaryotic and eukaryotic systems
Author: Tarar, Maryam Riaz
Awarding Body: University of London
Current Institution: University College London (University of London)
Date of Award: 1995
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The particulate form of the core antigen of hepatitis B virus (HBcAg), is highly immunogenic. It has been used as a carrier molecule, for expression and presentation of heterologous viral epitopes on the surface of hybrid core particles, in immunogenicity studies. The aim of this project was to produce a hybrid antigen comprising HBcAg and an immunogenic epitope of human cytomegalovirus (HCMV). A direct comparison was made of amino and carboxyl terminal fusions, by investigating the influence of position of the foreign epitope on antigenicity, immunogenicity and hybrid core particle formation. A part of the HCMV genome, encoding a neutralizing glycoprotein epitope gp58, was inserted at the amino terminus or fused to the truncated carboxyl terminus of HBcAg in separate constructs and expressed in a prokaryotic system (E.coli). At the same time, in order to express the same fusion proteins in a eukaryotic system, the baculovirus expression vector system (BEVS) was selected and as an initial control two recombinant baculoviruses, containing genes encoding HBcAg and hepatitis B surface antigen were isolated by dot blot hybridization. It was realized that work in BEVS would require more time than previously expected therefore further work was only carried out on the prokaryotic system. The HBcAg carboxyl terminal fusion (HBC 3 - 1 4 4 -HCMV) was expressed in high yields in E.coli and assembled into core like particles resembling native HBcAg. A similar fusion in the amino terminus of HBcAg (HCMV-HBC1-183) could not be purified or characterized immunologically, although it formed core like particles. HBc3-144-HCMV displayed HBc antigenicity but HCMV antigenicity could not be detected. Following immunization of rabbits with HBC3-144-HCMV, a high level of anti-HBc specific antibody was produced along with HCMV/gp58 specific antibody. The data presented here provide evidence that the HCMV/gp58 region can be used as a candidate immunogen for an HCMV subunit vaccine and that HBcAg can effectively present this foreign epitope joined to its carboxyl terminus, to the immune system.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available