title: Production of catalytically inactive BoNT/A1 holoprotein and comparison with BoNT/A1 subunit vaccines against toxin subtypes A1, A2, and A3.
abstract: A recombinant, catalytically inactive Clostridium botulinum neurotoxin A1 holoprotein (ciBoNT/A1 HP) was constructed by introducing amino acid substitutions H223A, E224A, and H227A in the active site to ablate proteolytic activity. ciBoNT/A1 HP was produced in the yeast Pichia pastoris and the purified product was evaluated as a vaccine candidate by comparison against recombinant BoNT/A1 LC, LC-belt, LC-H(n), and H(c) antigens and a LC-H(n)+H(c) combination in mouse potency and efficacy bioassays when challenged with BoNT/A subtypes /A1, /A2, and /A3. A single dose of ciBoNT/A1 HP provided equivalent or greater protective immunity, not only against the homologous toxin, but also against two distinct toxin subtypes with significant amino acid divergence. Only the LC-H(n)+H(c) combination provided comparable protection against /A1; however, it was less effective against subtypes /A2 and /A3. Differences in protective immunity diminished after multiple vaccinations with either ciBoNT/A1 HP or BoNT/A1 H(c), and the survival rates were more comparable at the toxin levels used to challenge.
authors.affiliation: United States Army Medical Research Institute for Infectious Diseases, 1425 Porter Street, Frederick, MD 21702, United States.
authors.forename: Robert P
authors.initials: RP
authors.lastname: Webb
authors.forename: Theresa J
authors.initials: TJ
authors.lastname: Smith
authors.forename: Patrick
authors.initials: P
authors.lastname: Wright
authors.forename: Jennifer
authors.initials: J
authors.lastname: Brown
authors.forename: Leonard A
authors.initials: LA
authors.lastname: Smith
mesh_terms: D000818:Animals; D001428:Bacterial Vaccines / Q000276:immunology*; D019274:Botulinum Toxins, Type A / Q000096:biosynthesis / Q000276:immunology* / Q000302:isolation & purification; D001906:Botulism / Q000276:immunology / Q000517:prevention & control*; D003014:Clostridium botulinum / Q000276:immunology; D007928:Lethal Dose 50; D051379:Mice; D010843:Pichia / Q000378:metabolism; D055550:Protein Stability; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification; D022223:Vaccines, Subunit / Q000276:immunology
chemical_list: D001428:Bacterial Vaccines; D011994:Recombinant Proteins; D022223:Vaccines, Subunit; D019274:Botulinum Toxins, Type A