The outer membrane (OM) proteins in gram-negative bacteria contain epitopes that are exposed on the cell surface (surface epitopes), and epitopes that are buried in the OM bilayer (buried epitopes). This proposal deals with the use of monoclonal antibodies (MAbs), already raised against the two classes of epitopes, to elucidate the role of major OM proteins in immune response to infection by Salmonella.
The specific aims of the project are to determine the extent of conservation of porin epitopes within the genus Salmonella, study the immunochemical structure of conserved domains, and evaluate the relative importance of surface and buried epitopes in humoral immune response to infection by Salmonella. the antigenic conservation of Salmonella porins will be determined by screening approximately 100 Salmonella serotypes by Western immunoblot against a panel of anti-(S. typhimurium) OmpD and OmpC MAbs. The immunochemical structure of the porin epitopes will be analyzed by Western immunoblot of antiporin MAbs against cyanogen bromide (CNBr) digests of porin monomers, and by amino acid and N- terminal sequence analyses of CNBr peptides. Finally, the relative importance of surface and buried epitopes in humoral immune response to infection will be studied by testing MAbs which recognize the two classes of epitopes for their ability to passively protect mice against infection by S. typhimurium. In the proposed investigation, murine salmonellosis which has many of the hallmarks of enteric fever will be used as a model system for the study of human typhoid disease induced by S. typhi. The long-term objective of this project is to elucidate the potential application of major OM proteins or peptides mimicking certain epitopes on these proteins as vaccines against S. typhi infections. Since all gram- negative bacteria probably contain similar OM protein structures, the proposed research is relevant to gram-negative bacterial pathogenesis in general. The project is also designed to train minority students who will participate in all phases of this research project. Students will review literature, formulate research ideas, design and carry out experiments using modern biomedical research techniques, and gain experience in writing and presentation of scientific papers at regional and national meetings.
