This research represents an effort to understand the molecular pathogenesis of intestinal diseases due to cytotoxin producing bacteria such as Shigella, cause of bacillary dysentery, and certain E. coli that causae watery diarrhea in infants (EPEC) and hemorrhagic colitis in children and adults (0157:H7). These organisms results in considerable morbidity in the U.S. and especially in the third world, and they are responsible for he majority of cases of enteric infection-related hemolytic-uremic syndrome, an often lethal complication, and also for most of the ORT-unresponsive diarrheal disease deaths in the third world. It is the goal of this research to understand how the toxin interacts with intestinal cells and its role in pathogenesis. Based on these data, we hope to develop better prophylactic, diagnostic and therapeutic approaches. These are three aims to the research: 1) to find out how many different distinct but biologically related toxins are produced, 2) to determine how many different tissue receptors there are, to characterize them, and to identify the biological process affected, and 3) to study the importance of the ability of toxin to inhibit protein synthesis in mediating the various toxin effects. Studies under Aim 1 will utilize physical, immunological, and functional criteria to assess the relatedness of purified toxins obtained from different organisms.
Aim 2 encompasses a variety of chemical, metabolic, and immunologic studies to define the fine structure and function of a glycolipid receptor we have described on rabbit intestinal cells, and to investigate whether or not a second glycoprotein receptor with different carbohydrate specificity exists for the cytotoxic activity of the toxin.
Aim 3 intends to produce of shigella:ricin hybrid A:B and B:A molecules, to determine whether or not the inactivation of 60S ribosomal subunits by the two toxins plays a role in enterotoxicity and whether or not B subunit targeting to a glycolipid receptor we have define (Gb3) is involved. We will also produce purified toxin for use in a collaboration with Dr. J. Robertus, University of Texas/San Antonio, to crystallize shigella toxin, determine its three dimensional X-ray structure, and compare it to ricin and attempt to define a common active site.
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