title: Immunogenicity of a Yersinia pestis vaccine antigen monomerized by circular permutation.
abstract: Caf1, a chaperone-usher protein from Yersinia pestis, is a major protective antigen in the development of subunit vaccines against plague. However, recombinant Caf1 forms polymers of indeterminate size. We report the conversion of Caf1 from a polymer to a monomer by circular permutation of the gene. Biophysical evaluation confirmed that the engineered Caf1 was a folded monomer. We compared the immunogenicity of the engineered monomer with polymeric Caf1 in antigen presentation assays to CD4 T-cell hybridomas in vitro, as well as in the induction of antibody responses and protection against subcutaneous challenge with Y. pestis in vivo. In C57BL/6 mice, for which the major H-2(b)-restricted immunodominant CD4 T-cell epitopes were intact in the engineered monomer, immunogenicity and protective efficacy were preserved, although antibody titers were decreased 10-fold. Disruption of an H-2(d)-restricted immunodominant CD4 T-cell epitope during circular permutation resulted in a compromised T-cell response, a low postvaccination antibody titer, and a lack of protection of BALB/c mice. The use of circular permutation in vaccine design has not been reported previously.
authors.affiliation: Institute for Cell and Molecular Biosciences, University of Newcastle upon Tyne, Framlington Place, Newcastle NE2 4HH, United Kingdom.
authors.forename: David A
authors.initials: DA
authors.lastname: Chalton
authors.forename: Julie A
authors.initials: JA
authors.lastname: Musson
authors.forename: Helen
authors.initials: H
authors.lastname: Flick-Smith
authors.forename: Nicola
authors.initials: N
authors.lastname: Walker
authors.forename: Alistair
authors.initials: A
authors.lastname: McGregor
authors.forename: Heather K
authors.initials: HK
authors.lastname: Lamb
authors.forename: E Diane
authors.initials: ED
authors.lastname: Williamson
authors.forename: Julie
authors.initials: J
authors.lastname: Miller
authors.forename: John H
authors.initials: JH
authors.lastname: Robinson
authors.forename: Jeremy H
authors.initials: JH
authors.lastname: Lakey
mesh_terms: D000818:Animals; D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001681:Biological Assay; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D006825:Hybridomas / Q000276:immunology; D016056:Immunodominant Epitopes / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D009154:Mutation; D010930:Plague / Q000517:prevention & control*; D010931:Plague Vaccine / Q000235:genetics / Q000276:immunology*; D011487:Protein Conformation; D015202:Protein Engineering; D017510:Protein Folding; D012996:Solutions / Q000737:chemistry; D015010:Yersinia pestis / Q000276:immunology*
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D016056:Immunodominant Epitopes; D010931:Plague Vaccine; D012996:Solutions; C086725:caf1 protein, Yersinia pestis