Use of the Enterobacterial Outer Membrane Protein PhoE in the Development of New Vaccines and DNA Probes

Abstract

PhoE protein is a major outer membrane protein of Escherichia coli. The polypeptide spans the membrane 16 times, thereby exposing 8 regions at the cell surface. Insertions in these regions did not affect the biogenesis of the protein. Therefore, we considered the possibility of using PhoE as a vector for the exposure of foreign antigenic determinants at the cell surface, with the ultimate goal of constructing new (live oral) vaccines. Via recombinant DNA techniques, B-cell epitopes of VP1 protein of foot-and-mouth-disease virus were inserted in the exposed regions of PhoE. The inserted epitopes were antigenic and immunogenic in the PhoE-associated conformation. Guinea pigs, immunized with such a hybrid protein were protected against viral challenge. Similarly, a T-cell epitope of the 65 kDa heat-shock protein of Mycobacterium tuberculosis remained antigenic and immunogenic in the PhoE-associated conformation, although recognition by the cells of the immune system was dependent on the amino acids, flanking the epitope. When the amino acid sequences of the PhoE proteins of different members of the family of Enterobacteriaceae are compared, the cell surface-exposed regions are hypervariable. Therefore, we considered the possibility that the DNA segments encoding these regions are species-specific. By using synthetic oligonucleotides corresponding to such DNA segments, primer couples for the specific detection and identification of different enterobacterial species, including Salmonella, by polymerase chain reactions have been developed.