title: Structural correlates of carrier protein recognition in tetanus toxoid-conjugated bacterial polysaccharide vaccines.
abstract: An analysis of structure-antibody recognition relationships in nine licenced polysaccharide-tetanus toxoid (TT) conjugate vaccines was performed. The panel of conjugates used included vaccine components to protect against disease caused by Haemophilus influenzae type b, Neisseria meningitidis groups A, C, W and Y and Streptococcus pneumoniae serotype 18C. Conformation and structural analysis included size exclusion chromatography with multi-angle light scattering to determine size, and intrinsic fluorescence spectroscopy and fluorescence quenching to evaluate the protein folding and exposure of Trp residues. A capture ELISA measured the recognition of TT epitopes in the conjugates, using four rat monoclonal antibodies: 2 localised to the HC domain, and 2 of which were holotoxoid conformation-dependent. The conjugates had a wide range of average molecular masses ranging from 1.8×10(6) g/mol to larger than 20×10(6) g/mol. The panel of conjugates were found to be well folded, and did not have spectral features typical of aggregated TT. A partial correlation was found between molecular mass and epitope recognition. Recognition of the epitopes either on the HC domain or the whole toxoid was not necessarily hampered by the size of the molecule. Correlation was also found between the accessibility of Trp side chains and polysaccharide loading, suggesting also that a higher level of conjugated PS does not necessarily interfere with toxoid accessibility. There were different levels of carrier protein Trp side-chain and epitope accessibility that were localised to the HC domain; these were related to the saccharide type, despite the conjugates being independently manufactured. These findings extend our understanding of the molecular basis for carrier protein recognition in TT conjugate vaccines.
authors: Division of Bacteriology, National Institute of Biological Standards and Control, Medicines and Healthcare Products Regulatory Agency, South Mimms, Potters Bar EN6 3QG, UK.
authors: Kay
authors: K
authors: Lockyer
authors: Division of Bacteriology, National Institute of Biological Standards and Control, Medicines and Healthcare Products Regulatory Agency, South Mimms, Potters Bar EN6 3QG, UK.
authors: Fang
authors: F
authors: Gao
authors: Faculty of Life Sciences, The University of Manchester, Manchester, UK.
authors: Jeremy P
authors: JP
authors: Derrick
authors: Division of Bacteriology, National Institute of Biological Standards and Control, Medicines and Healthcare Products Regulatory Agency, South Mimms, Potters Bar EN6 3QG, UK. Electronic address: barbara.bolgiano@nibsc.org.
authors: Barbara
authors: B
authors: Bolgiano
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002352:Carrier Proteins / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D008958:Models, Molecular; D011135:Polysaccharides, Bacterial / Q000276:immunology* / Q000302:isolation & purification; D017510:Protein Folding; D051381:Rats; D013745:Tetanus Toxoid / Q000737:chemistry / Q000276:immunology*; D018074:Vaccines, Conjugate / Q000737:chemistry* / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D011135:Polysaccharides, Bacterial; D013745:Tetanus Toxoid; D018074:Vaccines, Conjugate
keywords: Chromatography; Conjugate vaccine; Fluorescence quenching; Tetanus toxoid
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