Immunogenicity of aYersinia pestisVaccine 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.

The natural history and incidence of Yersinia pestis and prospects for vaccination

“…Progress is being made in this area since CD4 + Tcell epitopes in F1 and V have been identified in mice Shim et al, 2006) and an N-terminal H-2 d -restricted murine T-cell epitope in F1 has been shown to be essential for protection in BALB/c mice (Chalton et al, 2006). The significance of certain CD4 + Tcell epitopes in the F1 antigen for protection has been demonstrated in a study in which a monomeric stable circularly permutated F1 (cpF1) protein was created by relocating the N-terminal donor strand, via a flexible linker, to the C-terminus (Chalton et al, 2006). The cpF1 protein retained immunogenicity in both H-2 d and H-2 k mouse strains, but the circularization had the effect of disrupting the H-2 d restricted T-cell epitope (at primary sequence position 7-20) in the N-terminal donor strand, with loss of protective efficacy in BALB/c mice, but with no effect on protective efficacy in H-2 k mice.…”

Arbobakterien (über Arthropoden übertragbare Bakterien)

“…Die Abwehrreaktion im Menschen beruht auf Antikörpern und auf der schnellen Bildung von zytotoxischen T-Lymphozyten, die gegen die Virulenzfaktoren, die von dem 70-kb-Plasmid kodiert werden, gerichtet sind [82], ferner gegen Caf1, welches von dem 110-kb-Plasmid kodiert wird, einem als Chaperon wirkenden Protein [83]. NAT.…”

“…Ein Lebendimpfstoff mit einem in den Purinstoffwechsel eingreifenden, deletierten Protein von Francisella tularensis erzeugt in Mäusen eine Immunität, die sie gegen eine tödliche Dosis des Wildbakteriums schützt [90]. Eine Teilimmunität gegen Yersinia pestis mit einem Impfstoff, bestehend aus dem Cafl-Protein, lässt sich in Mäusen erzeugen [83].…”

Arbobacteria – Pathogens Transmittable by Arthropods

“…A live vaccine of F. tularensis with a deleted protein, which influences purin metabolism, creates immunity in mice protecting them from a deadly dose of the wild-type bacterium [90]. Partial immunity against Y. pestis with a vaccine consisting of Cafl protein can be created in mice [83].…”

“…This explains why Y. pestis strains with different degrees of pathogenicity occur in geographically restricted areas. The immune response in humans is based on antibodies and the rapid formation of cytotoxic T lymphocytes directed against virulence factors encoded by the 70 kb plasmid [82], and directed against Caf1 encoded by the 110 kb plasmid, a protein which acts as a chaperon [83].…”