Structural and biochemical analysis of the essential type III secretion system ATPase from gram-negative pathogenic bacteria

Gram-negative bacterial infections are a growing threat to humans, animals and plants. The rates of mortality and morbidity are further exasperated by increasingly antibiotic resistant forms of these pathogenic bacteria including strains of Yersinia, Shigella, Salmonella, Pseudomonas, Chlamydia and enteropathogenic E.coli (EPEC). Pathogenicity is acquired genetically via conjugation and defined by creation of pathogenicity islands that encode a complex macromolecular needle shape structure composed of more then 20 different proteins known as the type III secretion system (TTSS). Aided by chaperones, the TTSS delivers the virulence factors from the bacteria directly into the host cells using a largely uncharacterized mechanism. The energy for toxin insertion is provided by a highly conserved ATPase that is believed to be localized to the bacterial inner membrane. My project will focus on structure determination and characterization of the ATPase from EPEC (EscN) and complexes of the ATPase with virulence factors and/or their cognate chaperones. By providing the first 3D molecular structure and by understanding the mode of action of this type III ATPase, novel inhibitors can be designed and subsequently used as therapeutics against these common pathogens.