Viruses and plasmids control the production of several bacterial toxins. The heat-labile enterotoxin (LT) of Escherichia coli, a recently discovered LT-like toxin of E. coli, and diphtheria toxin (DT), the classical exotoxin of Corynebacterium diphtheriae, will be studied in this project. LT is directly involved in the pathogenesis of diarrhea in humans and animals; it is a model for control of toxinogenesis by plasmids. DT is directly involved in the pathogenesis of diphtheria and is a model for control of toxinogenesis by bacteriophages. In contrast, the role of LT-like toxin in disease has not yet been investigated. The long range goals of the proposed studies are to characterize these toxins and the molecular mechanisms that control their production and to apply this knowledge for control of infectious diseases in humans. A wide variety of biochemical, immunologic, genetic, and recombinant DNA methods will be used.
The specific aims for studying LT are to define the structure of specific antigenic determinants (epitopes) by using monoclonal anti-LT antibodies and synthetic oligopeptides related to LT and characterize mechanisms that regulate synthesis, processing and secretion of LT in E. coli.
The specific aims of the proposed studies of LT-like toxin are to purify the toxin, characterize its immunochemical properties and mode of action, clone and characterize the toxin structural gene(s), analyze the regulation of toxinogenesis, and determine the role of the toxin in pathogenesis of diarrheal diseases. The studies of LT and LT-like toxin will provide information that should be relevant for the design of synthetic oligopeptide vaccines and/or the construction of bacterial strains for use as living attenuated vaccines to protect against diarrheal diseases caused by E. coli.
The specific aims for studying DT are to characterize its interactions with artificial membrane vesicles and analyze the molecular basis for the role of iron in regulating toxinogenesis in C. diphtheriae. The studies with model membranes may shed light on the mechanism of translocation of DT across biological membranes. The studies on the role of iron will provide fundamental knowledge about regulation of toxinogenesis in C. diphtheriae. This knowledge may have practicalapplications for production of DT and related mutant proteins (CRMs) to be used for the preparation of conjugated vaccines or immunotoxins
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