The work proposed will focus primarily on the expression of five toxin types: enterotoxins A and B, alpha and beta toxins, and exfoliative toxin. The genetic and biosynthetic parameters governing these substances share a number of common features. Most prominently, they may belong to a new class of genetic elements characterized as hitch-hiking transposons. Secondarily, they are all transported by a similar mechanism which features cell wall compartmentalization of newly synthesized and processed toxin. The goals of the proposal are to provide information to build a model in Staphylococcus aureus describing the genetic and biosynthetic regulation of extracellular toxins, with particular emphasis on the food poisoning enterotoxins. This will be achieved by a blending of genetic and biochemical analyses designed to understand the movement of genetic material, the organization of the chromosome and collateral genetic elements, the biosynthesis of toxin molecules and the transport of these molecules to the extracellular medium. These aspects are important in understanding not only the appearance of so-called new virulent strains (as occurred in toxin shock syndrome) but also in designing strategies to defeat the establishment of infection. The basic procedures involve transduction and transformation to move genetic information, gene cloning to isolate genetic elements, in vitro protein synthesis to analyze the products of certain genetic elements, and immunochemical techniques to analyze specifically for the products of the foregoing.
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