Identification of Small-Molecule Inhibitors of Yersinia pestis Type III Secretion System YscN ATPase

• Published in: PloS one Vol. 6; no. 5; p. e19716
• Main Authors: Swietnicki, Wieslaw, Carmany, Daniel, Retford, Michael, Guelta, Mark, Dorsey, Russell, Bozue, Joel, Lee, Michael S., Olson, Mark A.
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 18.05.2011; Public Library of Science (PLoS)
• Subjects: Adenosine triphosphatase; Adenosine Triphosphatases - antagonists & inhibitors; Adenosine Triphosphatases - chemistry; Adenosine Triphosphatases - genetics; Adenosine Triphosphatases - secretion; Adenosine Triphosphate - metabolism; Analysis; Animals; Antibiotics; Antigens; ATPases; Bacteria; Bacterial Proteins - antagonists & inhibitors; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - secretion; Biochemistry; Biology; Bubonic plague; Burkholderia; Burkholderia mallei - drug effects; Burkholderia mallei - enzymology; Carrier Proteins - antagonists & inhibitors; Carrier Proteins - chemistry; Carrier Proteins - genetics; Carrier Proteins - secretion; Catalysis; Catalytic Domain; Cell culture; Cell Death - drug effects; Cells (Biology); Chemistry; Clonal deletion; Disease Models, Animal; E coli; Enzyme Inhibitors - analysis; Enzyme Inhibitors - pharmacology; Enzymes; Escherichia coli; Feasibility studies; Gene Deletion; Genes, Bacterial - genetics; Gram-negative bacteria; Health aspects; HeLa Cells; Homology; Humans; Hydrolysis - drug effects; Identification methods; Infections; Infectious diseases; Inhibitors; Inhibitory Concentration 50; Kinetics; Maltose-Binding Proteins - metabolism; Mammalian cells; Mammals; Medical research; Medicine; Mice; Models, Molecular; Pasteurella; Pathogens; Plague; Plague - microbiology; Protein Multimerization - drug effects; Protein transport; Proteins; Recombinant Fusion Proteins - metabolism; Salmonella; Secretion; Sequence Homology, Amino Acid; Small Molecule Libraries - analysis; Small Molecule Libraries - pharmacology; Therapeutics; Virulence; Virulence (Microbiology); Virulence factors; Yersinia; Yersinia enterocolitica; Yersinia pestis; Yersinia pestis - drug effects; Yersinia pestis - enzymology; Yersinia pestis - genetics; Yersinia pestis - pathogenicity; Zoonoses; United States > US; Maryland
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0019716

Yersinia pestis is a gram negative zoonotic pathogen responsible for causing bubonic and pneumonic plague in humans. The pathogen uses a type III secretion system (T3SS) to deliver virulence factors directly from bacterium into host mammalian cells. The system contains a single ATPase, YscN, necessary for delivery of virulence factors. In this work, we show that deletion of the catalytic domain of the yscN gene in Y. pestis CO92 attenuated the strain over three million-fold in the Swiss-Webster mouse model of bubonic plague. The result validates the YscN protein as a therapeutic target for plague. The catalytic domain of the YscN protein was made using recombinant methods and its ATPase activity was characterized in vitro. To identify candidate therapeutics, we tested computationally selected small molecules for inhibition of YscN ATPase activity. The best inhibitors had measured IC(50) values below 20 µM in an in vitro ATPase assay and were also found to inhibit the homologous BsaS protein from Burkholderia mallei animal-like T3SS at similar concentrations. Moreover, the compounds fully inhibited YopE secretion by attenuated Y. pestis in a bacterial cell culture and mammalian cells at µM concentrations. The data demonstrate the feasibility of targeting and inhibiting a critical protein transport ATPase of a bacterial virulence system. It is likely the same strategy could be applied to many other common human pathogens using type III secretion system, including enteropathogenic E. coli, Shigella flexneri, Salmonella typhimurium, and Burkholderia mallei/pseudomallei species.