title: A meningococcal vaccine antigen engineered to increase thermal stability and stabilize protective epitopes.
abstract: Factor H binding protein (FHbp) is part of two vaccines recently licensed for prevention of sepsis and meningitis caused by serogroup B meningococci. FHbp is classified in three phylogenic variant groups that have limited antigenic cross-reactivity, and FHbp variants in one of the groups have low thermal stability. In the present study, we replaced two amino acid residues, R130 and D133, in a stable FHbp variant with their counterparts (L and G) from a less stable variant. The single and double mutants decreased thermal stability of the amino- (N-) terminal domain compared with the wild-type protein as measured by scanning calorimetry. We introduced the converse substitutions, L130R and G133D, in a less stable wild-type FHbp variant, which increased the transition midpoint (Tm) for the N-terminal domain by 8 and 12 °C; together the substitutions increased the Tm by 21 °C. We determined the crystal structure of the double mutant FHbp to 1.6 Å resolution, which showed that R130 and D133 mediated multiple electrostatic interactions. Monoclonal antibodies specific for FHbp epitopes in the N-terminal domain had higher binding affinity for the recombinant double mutant by surface plasmon resonance and to the mutant expressed on meningococci by flow cytometry. The double mutant also had decreased binding of human complement Factor H, which in previous studies increased the protective antibody responses. The stabilized mutant FHbp thus has the potential to stabilize protective epitopes and increase the protective antibody responses to recombinant FHbp vaccines or native outer membrane vesicle vaccines with overexpressed FHbp.
authors.affiliation: Center for Immunobiology and Vaccine Development, Children's Hospital Oakland Research Institute, University of California San Francisco Benioff Children's Hospital Oakland, Oakland, CA 94609.
authors.forename: Monica
authors.initials: M
authors.lastname: Konar
authors.affiliation: Center for Immunobiology and Vaccine Development, Children's Hospital Oakland Research Institute, University of California San Francisco Benioff Children's Hospital Oakland, Oakland, CA 94609.
authors.forename: Rolando
authors.initials: R
authors.lastname: Pajon
authors.affiliation: Center for Immunobiology and Vaccine Development, Children's Hospital Oakland Research Institute, University of California San Francisco Benioff Children's Hospital Oakland, Oakland, CA 94609 pbeernink@chori.org.
authors.forename: Peter T
authors.initials: PT
authors.lastname: Beernink
mesh_terms: D019943:Amino Acid Substitution; D000942:Antigens, Bacterial / Q000737:chemistry* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D018360:Crystallography, X-Ray; D006358:Hot Temperature; D006801:Humans; D022401:Meningococcal Vaccines / Q000737:chemistry* / Q000235:genetics; D038541:Neisseria meningitidis, Serogroup B; D055550:Protein Stability
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D022401:Meningococcal Vaccines; C484699:factor H-binding protein, Neisseria meningitidis
keywords: Neisseria meningitidis; calorimetry; complement Factor H; crystal structure; site-specific mutagenesis