Structure and Function of Bacterial Toxins

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1. BACKGROUND: Bacteria use virulence factors as tools to aid in their colonization of host cells and in their quest for food. The mono-ADP-ribosyltransferase (mART) toxins is a large family of virulence factors produced by a wide range of bacterial pathogens. Many members are the principal causative agents in serious diseases in agriculture, including gastroenteritis and respiratory illnesses in livestock, pollinator diseases, and blight and scab diseases in crops. The mART toxins are enzymes that bind and cleave a key bond in NAD+ and covalently modify a critical target protein in the host. The covalent modification of the target protein (ADP-ribosylation) by mART toxins alters the host macromolecule function and can act as an “on” “off” switch for activity. 2. HYPOTHESIS: Structural and biochemical characterization of members of the mART toxin family, including their biological targets, will provide the basis for the eventual treatment of infections and diseases produced by the associated bacterial pathogens. 3. OBJECTIVES: The major objectives are (i) to employ a bioinformatic strategy to discover new members of the mART toxin family that will be characterized both structurally and functionally with a biochemical/biophysical approach; (ii) to identify their biological targets in a yeast model system to provide new insights on host-pathogen interactions; and (iii) to develop potent active-site inhibitors against these toxins as tools for further study and as the basis for future drug discovery and development. 4. NOVELTY AND SIGNIFICANCE: A compelling, innovative and alternative approach to antibiotic therapy is the anti-virulence strategy that involves targeting virulence-associated rather than survival/fitness-relevant traits in the offending pathogen. Anti-virulence agents are compounds that disarm bacteria without killing them, thereby allowing the infected host to use its normal immune system defenses to control or remove the infection. The ability to discover and design therapeutics against mART toxins lies in the depth of our understanding of these toxin/enzymes' structures and reaction mechanism. The suite of enzyme structures with matching substrates and inhibitors are beginning to bear fruit from our research efforts and will provide a virtual movie of the catalytic cycle for this important class of enzymes. These high-resolution data, combined with the focussed efforts of our team to improve and develop more potent inhibitors of these toxin-enzymes, may provide the basis for the future development of anti-virulence compounds for prophylactic treatment and prevention of bacterial diseases affecting agriculture and food.