SHIGELLA TOXIN; STRUCTURE & FUNCTION RELATIONSHIPS

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Shigellosis is a world wide disease producing significant morbidity and in developing countries, a leading cause of diarrheal disease mortality. The Shigella species which causes the severest disease, dysenteriae, produces a potent toxin molecule which inhibits protein synthesis. The toxin consists of one A chain and 5 B chains. A functional role of the B chain is to mediate toxin binding to the eukaryotic cell surface. The toxin receptors are glycolipids containing terminal Gal-alpha1->4Gal disaccharide. Shiga toxin is now considered to be the prototype of a family of toxins which have been called Shiga-like. Shiga and Shiga-like toxins are associated with many clinical manifestations including diarrhea, dysentery, hemorrhagic colitis, and the hemolytic uremic syndrome. This proposal is designed to study basic structure-function relationship of the toxin B chain. A monoclonal antibody which cross-reacts with the B chain of Shiga and Shiga-like toxin II will be characterized. Since these two toxins are thought to be immunologically distinct and since the two bind to an identical set of receptors, particular attention will be focused on whether this antibody is directed against the binding domain. A monoclonal antibody and a glycoprotein with Gal-alpha1->4Gal terminal disaccharide, P1-glycoprotein, will be used as tools to select bacterial strains which produce B subunits with binding domain alterations. The characterization of these mutants will provide an understanding of the regions of the B chain involved in receptor recognition. This information will be useful for the future design of safe and effective vaccines and for the potential design of therapeutic intervention regimens. Additionally the biochemical and genetic characterization of a material which cross- reacts with antibody against Shiga toxin will be undertaken with particular emphasis on revealing the role this material may play in pathogenesis. Finally, the uncharacterized cytotoxins produced by other Gram-negative organisms will be analyzed for their Shiga-like properties. Information as to whether other pathogenic bacteria produce Shiga-like toxins will aid in our understanding of the spectrum of disease associated with toxin.