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Infect. Immun. doi:10.1128/IAI.00437-06
Copyright (c) 2006, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Immunogenicity of a Yersinia pestis vaccine antigen monomerised by circular permutation

Institute for Cell and Molecular Biosciences, University of Newcastle upon Tyne, Framlington Place, Newcastle, NE2 4HH, U.K; Musculoskeletal Research Group, Clinical Medical Sciences, University of Newcastle upon Tyne, Framlington Place, Newcastle, NE2 4HH, U.K.; and Defence Science and Technology Laboratory, Porton Down, Salisbury, Wiltshire, SP4 0JQ, UK

^* To whom correspondence should be addressed. Email: j.h.lakey{at}ncl.ac.uk.

	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 was 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 post-vaccination antibody titer and lack of protection of BALB/c mice. Use of circular permutation in vaccine design has not been reported previously.