title: A Mutant Library Approach to Identify Improved Meningococcal Factor H Binding Protein Vaccine Antigens.
abstract: Factor H binding protein (FHbp) is a virulence factor used by meningococci to evade the host complement system. FHbp elicits bactericidal antibodies in humans and is part of two recently licensed vaccines. Using human complement Factor H (FH) transgenic mice, we previously showed that binding of FH decreased the protective antibody responses to FHbp vaccination. Therefore, in the present study we devised a library-based method to identify mutant FHbp antigens with very low binding of FH. Using an FHbp sequence variant in one of the two licensed vaccines, we displayed an error-prone PCR mutant FHbp library on the surface of Escherichia coli. We used fluorescence-activated cell sorting to isolate FHbp mutants with very low binding of human FH and preserved binding of control anti-FHbp monoclonal antibodies. We sequenced the gene encoding FHbp from selected clones and introduced the mutations into a soluble FHbp construct. Using this approach, we identified several new mutant FHbp vaccine antigens that had very low binding of FH as measured by ELISA and surface plasmon resonance. The new mutant FHbp antigens elicited protective antibody responses in human FH transgenic mice that were up to 20-fold higher than those elicited by the wild-type FHbp antigen. This approach offers the potential to discover mutant antigens that might not be predictable even with protein structural information and potentially can be applied to other microbial vaccine antigens that bind host proteins.
authors.affiliation: Center for Immunobiology and Vaccine Development, Children's Hospital Oakland Research Institute, UCSF Benioff Children's Hospital Oakland, Oakland, CA, United States of America.
authors.forename: Monica
authors.initials: M
authors.lastname: Konar
authors.affiliation: Center for Immunobiology and Vaccine Development, Children's Hospital Oakland Research Institute, UCSF Benioff Children's Hospital Oakland, Oakland, CA, United States of America.
authors.forename: Raffaella
authors.initials: R
authors.lastname: Rossi
authors.affiliation: Center for Immunobiology and Vaccine Development, Children's Hospital Oakland Research Institute, UCSF Benioff Children's Hospital Oakland, Oakland, CA, United States of America; Biology Department, Mills College, Oakland, CA, United States of America.
authors.forename: Helen
authors.initials: H
authors.lastname: Walter
authors.affiliation: Center for Immunobiology and Vaccine Development, Children's Hospital Oakland Research Institute, UCSF Benioff Children's Hospital Oakland, Oakland, CA, United States of America.
authors.forename: Rolando
authors.initials: R
authors.lastname: Pajon
authors.affiliation: Center for Immunobiology and Vaccine Development, Children's Hospital Oakland Research Institute, UCSF Benioff Children's Hospital Oakland, Oakland, CA, United States of America.
authors.forename: Peter T
authors.initials: PT
authors.lastname: Beernink
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000917:Antibody Formation / Q000276:immunology; D000942:Antigens, Bacterial / Q000276:immunology*; D001426:Bacterial Proteins / Q000276:immunology*; D002152:Calorimetry, Differential Scanning; D017242:Complement Factor H / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism; D005260:Female; D015723:Gene Library; D006801:Humans; D055767:Immobilized Proteins / Q000378:metabolism; D022401:Meningococcal Vaccines / Q000276:immunology*; D008822:Mice, Transgenic; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D055550:Protein Stability; D011994:Recombinant Proteins / Q000378:metabolism; D044967:Serum / Q000378:metabolism; D020349:Surface Plasmon Resonance
chemical_list: D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C493495:CFH protein, human; D055767:Immobilized Proteins; D022401:Meningococcal Vaccines; D050505:Mutant Proteins; D011994:Recombinant Proteins; D017242:Complement Factor H