USE OF THE ENTEROBACTERIAL OUTER-MEMBRANE PROTEIN PHOE IN THE DEVELOPMENT OF NEW VACCINES AND DNA PROBES

PhoE protein is a major outer membrane protein of Escherichia coli. Th e polypeptide spans the membrane 16 times, thereby exposing 8 regions at the cell surface. Insertions in these regions did not affect the bi ogenesis of the protein. Therefore, we considered the possibility of u sing PhoE as a vector for the exposure of foreign antigenic determinan ts at the cell surface, with the ultimate goal of constructing new (li ve oral) vaccines. Via recombinant DNA techniques, B-cell epitopes of VP1 protein of foot-and-mouth-disease virus were inserted in the expos ed regions of PhoE. The inserted epitopes were antigenic and immunogen ic in the PhoE-associated conformation. Guinea pigs, immunized with su ch a hybrid protein were protected against viral challenge. Similarly, a T-cell epitope of the 65 kDa heat-shock protein of Mycobacterium tu berculosis remained antigenic and immunogenic in the PhoE-associated c onformation, although recognition by the cells of the immune system wa s dependent on the amino acids, flanking the epitope. When the amino a cid sequences of the PhoE proteins of different members of the family of Enterobacteriaceae are compared, the cell surface-exposed regions a re hypervariable. Therefore, we considered the possibility that the DN A segments encoding these regions are species-specific. By using synth etic oligonucleotides corresponding to such DNA segments, primer coupl es for the specific detection and identification of different enteroba cterial species, including Salmonella, by polymerase chain reactions h ave been developed.