Martino, Angela (2013) A new serogroup B meningococcal vaccine : biophysical characterisation of its protein components. PhD thesis, University of Warwick.

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Abstract

Meningococcal infection due to serogroup B disease is the leading cause of meningitis across Europe, particularly in infants. Prevention through vaccination will soon be possible utilising a new multicomponent vaccine (4CMenB or Bexsero®) principally directed against serogroup B Neisseria meningitidis. In addition to OMVs, the vaccine contains three novel recombinant proteins discovered by reverse vaccinology. Studies were performed on the biophysical characteristics of these antigens to better understand their structural properties in solution and aspects of their immunogenicity. Initially, the antibody responses to the vaccine components were assessed in a low-dose mouse model. This extreme immunogenicity model was chosen to highlight any subtle differences in the immune response to individual antigens which may have been overlooked by previously high-dose immunisation studies.
The recombinant vaccine components are synthetic homologues of proteins naturally present on the bacterial cell and changes in their structure can induce loss of functional responses. By deliberately stressing the antigens to lose functional immune responses, it was possible to study the structural changes at the molecular level. Using molecular modeling and protein-specific monoclonal and polyclonal antibody binding in parallel with UV circular dichroism (CD) spectroscopy, it was possible to assess at which point changes in the structure of proteins affected their ability to induce functional antibody responses in vivo. The two fusion proteins present unique biophysical characteristics derived from their component parts. As a result, they were shown to possess distinctive profiles by CD. When combined with fluorescence unfolding studies, in which the tryptophan residues were used as reporters for integrity, CD uncovered significant structural differences among the vaccine antigens. The NHBA-FP comprises two domains having different thermodynamics while in the fHbp-fusion both halves cooperate in the unfolding process. The choice of including a β-barrel protein in the chimeric constructs proved successful as both the NHBA and fHbp exhibited excellent solvent accessibility profiles due to the numerous inter- and intra-strand hydrogen bonds stabilising the structure. This had a direct effect on their immunogenicity which could only be adversely affected by prolonged harsh treatments. Maintenance of the α-helical profile was found to be critical for the immunogenicity of NadAΔ351–405 component as was the maintenance of its trimeric organisation.
Overall, results confirmed the requirement and importance of the three additional protein antigens to the bactericidal response, even when directed against the homologous OMV bacterial strain. Structural studies confirmed the quality of the vaccines antigens and supported their critical contribution to the vaccine.

Item Type:	Thesis (PhD)
Subjects:	Q Science > QR Microbiology R Medicine > RC Internal medicine R Medicine > RM Therapeutics. Pharmacology
Library of Congress Subject Headings (LCSH):	Meningitis, Cerebrospinal -- Treatment -- Research, Neisseria meningitidis, Bacterial vaccines -- Research, Physical biochemistry, Protein drugs
Official Date:	April 2013
Institution:	University of Warwick
Theses Department:	Molecular Organisation and Assembly in Cells; Department of Chemistry
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Rodger, Alison
Sponsors:	Meningitis Research Foundation (0602.0)
Extent:	xvii, 195 leaves : charts, illustrations.
Language:	eng

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